The reverse transcriptase (RT) model of immunoglobulin (Ig) somatic hypermutation (SHM) has recei... more The reverse transcriptase (RT) model of immunoglobulin (Ig) somatic hypermutation (SHM) has received insufficient scientific attention. This is understandable given that DNA deamination mediated by activationinduced deaminase (AID), the initiating step of Ig SHM, has dominated experiments since 2002. We summarise some key history of the RT Ig SHM model dating to 1987. For example, it is now established that DNA polymerase η, the sole DNA repair polymerase involved in post-replication short-patch repair, is an efficient cellular RT. This implies that it is potentially able to initiate target site reverse transcription by RNA-directed DNA repair at AID-induced lesions. Recently, DNA polymerase θ has also been shown to be an efficient cellular RT. Since DNA polymerase θ plays no significant role in Ig SHM, it could serve a similar RNA-dependent DNA polymerase role as DNA polymerase η at non-Ig loci in the putative RNA-templated nucleotide excision repair of bulky adducts and other mutagenic lesions on the transcribed strand. A major yet still poorly recognised consequence of the proposed RT process in Ig SHM is the generation of significant and characteristic strandbiased mutation signatures at both deoxyadenosine/deoxythymidine and deoxyguanosine/deoxycytidine base pairs. In this historical perspective, we highlight how diagnostic strand-biased mutation signatures are detected in vivo during SHM at both Ig loci in germinal centre B lymphocytes and non-Ig loci in cancer genomes. These strand-biased signatures have been significantly obscured by technical issues created by improper use of the polymerase chain reaction technique. A heightened awareness of this fact should contribute to better data interpretation and somatic mutation pattern recognition both at Ig and non-Ig loci.
Understanding the genesis of the block haplotype structure of the genome is a major challenge. Wi... more Understanding the genesis of the block haplotype structure of the genome is a major challenge. With the completion of the sequencing of the Human Genome and the initiation of the HapMap project the concept that the chromosomes of the mammalian genome are a mosaic, or patchwork, of conserved extended block haplotype sequences is now accepted by the mainstream genomics research community. Ancestral Haplotypes (AHs) can be viewed as a recombined string of smaller Polymorphic Frozen Blocks (PFBs). How have such variant extended DNA sequence tracts emerged in evolution? Here the relevant literature on the problem is reviewed from various fields of molecular and cell biology particularly molecular immunology and comparative and functional genomics. Based on our synthesis we then advance a testable molecular and cellular model. A critical part of the analysis concerns the origin of the strand biased mutation signatures in the transcribed regions of the human and higher primate genome, A-to-G versus Tto-C (ratio ~1.5 fold) and C-toT versus G-to-A (≥1.5 fold). A comparison and evaluation of the current state of the fields of immunoglobulin Somatic Hypermutation (SHM) and Transcription-Coupled DNA Repair (TCR) focused on how mutations in newly synthesized RNA might be copied back to DNA thus accounting for some of the genome-wide strand biases (e.g. the A-to-G vs T-to-C component of the strand biased spectrum). We hypothesize that the genesis of PFBs and extended AHs occurs during mutagenic episodes in evolution (e.g. retroviral infections) and that many of the critical DNA sequence diversifying events occur first at the RNA level e.g. recombination between RNA strings resulting in tandem and dispersed RNA duplications (retroduplications), RNA mutations via adenosine-to-inosine pre-mRNA editing events as well as error prone RNA synthesis. These are then copied back into DNA by a cellular reverse transcription (RT) process (also likely to be error-prone) we have called "RTmediated long DNA conversion" (RT-LDC). Finally we suggest that all these activities and others can be envisaged as being brought physically under the umbrella of special sites in the nucleus involved in transcription known as "Transcription Factories" (TF).
Recent innovations in microelectronics and advances in cryptography are driving the appearance of... more Recent innovations in microelectronics and advances in cryptography are driving the appearance of a new generation of smart cards with wider applications; this has important repercussions for our society in the coming years. Essentially, these breakthroughs include built-in microprocessors capable of generating cryptographic transactions (e.g.,Jelectronic blinded signatures, digital pseudonyms, and digital credentials), developments toward a single electronic card offering multi-access to services such as transport, telecommunications, health, financial, and entertainment (Universal Access Services), and incorporation of personal identification technologies such as voice, eye, or skin pattern recognition. For example, by using electronic representatives or cryptographic blinded signatures, a smart card can be used for multi transactions across different organizations and under different generated pseudonyms. These pseudonyms are capable of recognizing an individual unambiguously, while none of her records can be linked [1]. Moreover, tamper-proof electronic observers would make smart cards a very attractive technology for high-security based applications, such as those in the health care field. New trends in smart card technology offer excellent privacy and confidentiality safeguards. Therefore, smart cards constitute a promising technology for the health sector in Australia and other countries around the world in their pursuit of technology to support the delivery of quality care services. This paper addresses the main issues and the key design criteria which may be of strategic importance to the success of future smart card technology in the health care sector.
Evidence already exists that the activation-induced deaminase (AID)/APOBEC family constitutes a s... more Evidence already exists that the activation-induced deaminase (AID)/APOBEC family constitutes a set of differentially expressed enzymes capable of deaminating cytosines (C to U) in singlestranded DNA (ssDNA) and that they are potentially powerful mutagens. The mutagenic processes involved are believed to be activated in many nonlymphoid tissue typesdfor example, initiating some cancers and/or leading to further somatic mutagenesis. To investigate the extent that codon context might be important in influencing the likely location of TP53 mutations in breast cancer, the codon-bias patterns resulting from the ssDNA target specificities of cytidine deaminases of the AID/APOBEC family were analyzed. The data indicate that codon context strongly influences the likely location of mutations at motifs for AID/APOBEC1/APOBEC3G, and at WA sites. An unexpected finding is a highly significant preference for transitions of cytosine to occur at the first nucleotide position and for transitions of guanosine to occur at the second nucleotide position in the mutated codon (read 3 0 to 5 0). Thus, the mechanisms involved appear to be sensitive to codon reading frames and to have an intrinsic ability to differentiate between the cytosines on the nontranscribed strand and those on the transcribed strand in the context of an open "transcription bubble."
Mutation Research: Fundamental And Molecular Mechanisms Of Mutagenesis, Jul 1, 2018
Cytosine and adenosine deamination events (DNA, RNA substrates) account for most codon-context Ta... more Cytosine and adenosine deamination events (DNA, RNA substrates) account for most codon-context Targeted Somatic Mutation (TSM) patterns observed in immunoglobulin (Ig) somatic hypermutation (SHM), and in cancer exomes following Ig-SHM-like responses. TSM refers to the process of somatic mutagenesis involving deamination events that results on a dominant type of mutation (e.g., C-toT), and co-incident at a particular motif (e.g., WRC), and preferentially targeting the first, second or third nucleotide position within the mutated codon (e.g. MC1, MC2 or MC3, read 5-prime to 3-prime). It is now widely accepted that if left uncorrected, the accumulation of uncorrected TSMs involving the deaminases, may lead to a diagnosis of cancer or other degenerative disease. Our hypothesis is that many missense, nonsense and synonymous single nucleotide polymorphisms (SNPs) associated with clinically significant diseases may have arisen in the population by similar highly targeted deamination events. The OMIM database was searched for disease-associated SNPs on the X chromosome, and for all chromosomes. The nucleotide substitution patterns for disease-associated SNPs were analyzed by the TSM method to identify the likely deaminase source for C-to-U
This paper simply links the findings of Zheng, Lorenzo and Beal (2017) to our previous work on st... more This paper simply links the findings of Zheng, Lorenzo and Beal (2017) to our previous work on strand biased and codon-context mutation signatures in B lymphocytes (Ig SHM) and codon-contexted exomewide point mutation patterns in cancer genomes. We conclude that in vivo the A-to-I DNA editing component at RNA: DNA hybrids occurring in Transcription Bubbles, while important, is of far lower Ato-I editing efficiency than in dsRNA substrates (as shown in Zheng et al 2017). The extreme strand biased mutation patterns documented by us in vivo should be logically rationalized by the predicted sequential steps of the RNA/RT-based mechanism. Abbreviations used in this paper: Aag, alkyladenine DNA glycosylase; ADAR, Adenosine Deaminase that acts on RNA; ADAT, Adenosine Deaminase that acts on tRNA; AID, activation induced cytidine deaminase, a APOBEC family member, initiating via C-to-U lesions in ssDNA of class switch recombination (CSR) and somatic hypermutation (SHM) processes at somatically rearranged Ig V(D)J gene loci, and known to activate cytidine mutagenic deamination during transcription in other somatic tissues, particularly in cancer; APOBEC family, generic abbreviation for the deoxyribonucleic acid, or dCto-dU, deaminase family (APOBEC3 A, B, C, D, F, G, H) similar in DNA sequence to the "apolipoprotein B RNA editor" APOBEC1, and known to activate mutagenic cytidine deamination during transcription in somatic tissues, particularly in cancer; AP, an Abasic, or apurinic/apyrimidinic, site; APE, AP endonuclease; A-to-I, adenosine-toinosine RNA editing; BER, base excision repair; Deaminase, catalytic domain in ADAR and AID/APOBEC enzymes; DSB, double strand DNA breaks; Ig-SHM-like response, strand-biased somatic mutation patterns similar to that observed in Ig SHM; MMR, mismatch repair; Motif, 4 to 6 nucleotide (N) sequence defining specificity of deaminase targeting; MSH2-MSH6, MutSa heterodimer recognising mispaired bases in DNA duplex; N, any nucleotide; NTS, the non-transcribed, or "Top", strand; NGS, Next Generation Sequencing; Pol-h or DNA polymerase-h (eta); R, Adenosine (A) or Guanine (G) , purines; RNA Pol II, RNA Polymerase II; RT, reverse transcriptase; RT-Pol-h, reverse transcriptase activity displayed by Pol-h; S, strong base pair involving Cytosine (C) or Guanine (G); SHM, somatic hypermutation; T, Thymine; TS, the transcribed, or "Bottom", strand, in context of a Transcription Bubble; TSM, targeted somatic mutations : the process of targeting C and A nucleotides for deamination in actively transcribed genes that results in a dominant type of mutation caused by a DBD or Inf-DBD at a particular codon position; TSRT, target site reverse transcription; U, uracil; UNG, uracyl DNA glycosylase involved in BER at dU sites in DNA resulting in either an Abasic site (AP) or APE-mediated ssDNA nicks (above); UTR, untranslated regions in the upstream (5') and downstream (3') regulatory regions of protein coding genes; V(D)J, generic symbol for a rearranged immunoglobulin (or T cell receptor, TCR) variable region gene in the
Targeted Somatic Mutation (TSM) analysis for disease-associated SNPs on all genes that are: I X C... more Targeted Somatic Mutation (TSM) analysis for disease-associated SNPs on all genes that are: I X Chromosome OMIM SNPs; and, II All chromosome OMIM SNPs in Clinvar database.
This paper reports the results of our initial analysis of APOBEC and ADAR deaminase-mediated muta... more This paper reports the results of our initial analysis of APOBEC and ADAR deaminase-mediated mutation signature patterns in complete COVID-19 genomes from informative locations and times in China, USA and Spain in the 2019-2020 pandemic. We have identified a unique set of 'new' putative coordinated Riboswitches in COVID-19 genomes not previously identified, and likely generating variants of the known common strain Haplotypes now in circulation. The results reveal that COVID-19 diversifies using switching of RNA Haplotypes with minimal alteration to protein structure (the normal targets for B and T cells in conventional vaccine development). The deaminase-driven RNA Haplotypes are most likely aligned with RNA secondary structures as several studies already highlight how Riboswitches alter he ability of RNA to fold into intricate three-dimensional structures allowing them to execute their diverse cellular functions. The same functional outcomes are expected for viruses, partic...
In hepatocellular cancer (HCC) there is an over expression of the RNA editing enzyme ADAR1. Furth... more In hepatocellular cancer (HCC) there is an over expression of the RNA editing enzyme ADAR1. Further, the prominent genomic somatic mutation signature in HCC is almost exclusively focused on mutations at A:T base pairs where A-to-G mutations far exceed T-to-C mutations (when read on the non-transcribed strand). A clear mechanism for this extreme transcriptional strand biased mutation signature, putatively associated with over expression of ADAR1 deaminase, is yet to be explicitly demonstrated. The standard description of this strand bias has been nominally called “Transcription Coupled Damage” (TCD) to distinguish it from more conventional “Transcription Coupled Repair” (TCR). We show that the TCD description does not satisfy all features of the molecular evidence. The conventional view is that ADAR1 is thought to target adenosines at WA-sites for editing to inosine (I) in double stranded RNA stem-loop structures in transcripts. Here we show that the totality of the molecular and cel...
This chapter addresses the molecular mechanism and source of the numerous mutagenic changes that ... more This chapter addresses the molecular mechanism and source of the numerous mutagenic changes that genomes, particularly mammalian genomes, can undergo during normal development and in diseased states. The central role of pathogen and other disease-inducing innate immunity via the action of the cytosine (AID/APOBEC) and adenosine (ADAR) deaminases is reviewed in some depth. The general and universal nature of deaminase-mediated mutagenesis is an important key to understanding cosmic genetic evolutionary processes.
Cytosine and adenosine deamination events (DNA, RNA substrates) account for most codon-context Ta... more Cytosine and adenosine deamination events (DNA, RNA substrates) account for most codon-context Targeted Somatic Mutation (TSM) patterns observed in immunoglobulin (Ig) somatic hypermutation (SHM), and in cancer exomes following Ig-SHM-like responses. TSM refers to the process of somatic mutagenesis involving deamination events that results on a dominant type of mutation (e.g., C-to-T), and co-incident at a particular motif (e.g., WRC), and preferentially targeting the first, second or third nucleotide position within the mutated codon (e.g. MC1, MC2 or MC3, read 5-prime to 3-prime). It is now widely accepted that if left uncorrected, the accumulation of uncorrected TSMs involving the deaminases, may lead to a diagnosis of cancer or other degenerative disease. Our hypothesis is that many missense, nonsense and synonymous single nucleotide polymorphisms (SNPs) associated with clinically significant diseases may have arisen in the population by similar highly targeted deamination event...
Evidence already exists that the activation-induced cytidine deaminase (AID/APOBEC) and the adeno... more Evidence already exists that the activation-induced cytidine deaminase (AID/APOBEC) and the adenosine deaminase (ADAR) families of enzymes are implicated as powerful mutagens in oncogenic processes in many somatic tissues. Each deaminase is identified by the DNA or RNA nucleotide sequence ("motif") surrounding the nucleotide targeted for deamination. The primary objective of this study is to develop an in silico approach to identify nucleotide sequence changes of the target motifs of key deaminases during oncogenesis. If successful, a secondary objective is to investigate if such changes are associated with disease progression indicators that include disease stage and progression-free survival time. Using a discovery cohort of 194 high-grade serous ovarian adenocarcinoma (HGS-OvCa) exomes, the results confirm the ability of the novel in silico approach used to identify changes in the preferred target motifs for AID, APOBEC3G, APOBEC3B, and ADAR1 during oncogenesis. Using t...
There are strong indications that now is an appropriate time for introducing a health insurance s... more There are strong indications that now is an appropriate time for introducing a health insurance smart card in Australia. Reviews of health implementations in Europe to date verify that a critical mass of projects exists and that technological requirements have been satisfied. Costs of card readers have declined to a desirable level, the security features of smart card may be employed to overcome confidentiality and privacy concerns, and opportunities exist to improve the efficiency and cost effectiveness of health care delivery and administration on a national level. However for a successful health insurance smart card system to be implemented in Australia, unique socio-cultural and economic contexts must be incorporated into the systems design. These include implementation on a national scale to ensure coherency, compatibility, and interoperability between insurance and healthcare providers; modifying existing health information systems to migrate to the smart card network; convincing medical practitioners to adopt guidelines on electronic medical records; and ensuring privacy and confidentiality of patient records through a mixture of legislative, administrative, and technical controls. When these criteria are satisfied in the smart card insurance design, the opportunity for this technology to provide customised benefits, improved efficiency, and a speedier delivery of services in the current socioeconomic climate will be realised.
This thesis breaks new ground by providing the first detailed study of smart card innovation duri... more This thesis breaks new ground by providing the first detailed study of smart card innovation during its first twenty years (1974-1996). The overall aim is to apply sociotechnical principles to further our understanding of the innovation process as it relates to smart card technology. By using a sociotechnical framework, this study also seeks to illustrate the limitations of conventional innovation
Evidence already exists that the activation-induced deaminase (AID)/APOBEC family constitutes a s... more Evidence already exists that the activation-induced deaminase (AID)/APOBEC family constitutes a set of differentially expressed enzymes capable of deaminating cytosines (C to U) in single-stranded DNA (ssDNA) and that they are potentially powerful mutagens. The mutagenic processes involved are believed to be activated in many nonlymphoid tissue types-for example, initiating some cancers and/or leading to further somatic mutagenesis. To investigate the extent that codon context might be important in influencing the likely location of TP53 mutations in breast cancer, the codon-bias patterns resulting from the ssDNA target specificities of cytidine deaminases of the AID/APOBEC family were analyzed. The data indicate that codon context strongly influences the likely location of mutations at motifs for AID/APOBEC1/APOBEC3G, and at WA sites. An unexpected finding is a highly significant preference for transitions of cytosine to occur at the first nucleotide position and for transitions of...
Activation-induced cytidine deaminase (AID) initiates Phase I somatic hypermutation (SHM) of anti... more Activation-induced cytidine deaminase (AID) initiates Phase I somatic hypermutation (SHM) of antibody genes by deaminating deoxy-cytosine to deoxy-uracil (C-to-U). These lesions trigger Phase II, a poorly understood process of error-prone repair targeting AT pairs by DNA polymerase (Pol). Since Pol is also a reverse transcriptase, Phase II could involve copying off RNA as well as DNA templates. We explore this idea further since in an RNAbased pathway it is conceivable that adenosine-to-inosine (A-to-I) RNA editing causes Ato-G transitions since I like G pairs with C. Adenosine deaminases (ADARs) are known to preferentially edit A nucleotides that are preceded by an A or U (W) in double-stranded RNA substrates. On this assumption and using a theoretical bioinformatics approach we show that a significant and specific correlation (P < 0.002) exists between the frequency of WA-to-WG mutations and the number of mRNA hairpins that could potentially form at the mutation site. This implies roles for both RNA editing and reverse transcription during SHM in vivo and suggests definitive genetic experiments targeting the appropriate ADAR1 isoform (␥INF-ADAR1) and/or Ig pre-mRNA templates.
... systems design: an Australian case study JOAN COOPER, NILAY GENCTURK and ROBYN A. LINDLEY Dep... more ... systems design: an Australian case study JOAN COOPER, NILAY GENCTURK and ROBYN A. LINDLEY Department of Information and Communication Technology (IACT), The University of Wollongong, NorthŪelds Avenue, Wollongong, 2500 Australia Abstract. ...
The reverse transcriptase (RT) model of immunoglobulin (Ig) somatic hypermutation (SHM) has recei... more The reverse transcriptase (RT) model of immunoglobulin (Ig) somatic hypermutation (SHM) has received insufficient scientific attention. This is understandable given that DNA deamination mediated by activationinduced deaminase (AID), the initiating step of Ig SHM, has dominated experiments since 2002. We summarise some key history of the RT Ig SHM model dating to 1987. For example, it is now established that DNA polymerase η, the sole DNA repair polymerase involved in post-replication short-patch repair, is an efficient cellular RT. This implies that it is potentially able to initiate target site reverse transcription by RNA-directed DNA repair at AID-induced lesions. Recently, DNA polymerase θ has also been shown to be an efficient cellular RT. Since DNA polymerase θ plays no significant role in Ig SHM, it could serve a similar RNA-dependent DNA polymerase role as DNA polymerase η at non-Ig loci in the putative RNA-templated nucleotide excision repair of bulky adducts and other mutagenic lesions on the transcribed strand. A major yet still poorly recognised consequence of the proposed RT process in Ig SHM is the generation of significant and characteristic strandbiased mutation signatures at both deoxyadenosine/deoxythymidine and deoxyguanosine/deoxycytidine base pairs. In this historical perspective, we highlight how diagnostic strand-biased mutation signatures are detected in vivo during SHM at both Ig loci in germinal centre B lymphocytes and non-Ig loci in cancer genomes. These strand-biased signatures have been significantly obscured by technical issues created by improper use of the polymerase chain reaction technique. A heightened awareness of this fact should contribute to better data interpretation and somatic mutation pattern recognition both at Ig and non-Ig loci.
Understanding the genesis of the block haplotype structure of the genome is a major challenge. Wi... more Understanding the genesis of the block haplotype structure of the genome is a major challenge. With the completion of the sequencing of the Human Genome and the initiation of the HapMap project the concept that the chromosomes of the mammalian genome are a mosaic, or patchwork, of conserved extended block haplotype sequences is now accepted by the mainstream genomics research community. Ancestral Haplotypes (AHs) can be viewed as a recombined string of smaller Polymorphic Frozen Blocks (PFBs). How have such variant extended DNA sequence tracts emerged in evolution? Here the relevant literature on the problem is reviewed from various fields of molecular and cell biology particularly molecular immunology and comparative and functional genomics. Based on our synthesis we then advance a testable molecular and cellular model. A critical part of the analysis concerns the origin of the strand biased mutation signatures in the transcribed regions of the human and higher primate genome, A-to-G versus Tto-C (ratio ~1.5 fold) and C-toT versus G-to-A (≥1.5 fold). A comparison and evaluation of the current state of the fields of immunoglobulin Somatic Hypermutation (SHM) and Transcription-Coupled DNA Repair (TCR) focused on how mutations in newly synthesized RNA might be copied back to DNA thus accounting for some of the genome-wide strand biases (e.g. the A-to-G vs T-to-C component of the strand biased spectrum). We hypothesize that the genesis of PFBs and extended AHs occurs during mutagenic episodes in evolution (e.g. retroviral infections) and that many of the critical DNA sequence diversifying events occur first at the RNA level e.g. recombination between RNA strings resulting in tandem and dispersed RNA duplications (retroduplications), RNA mutations via adenosine-to-inosine pre-mRNA editing events as well as error prone RNA synthesis. These are then copied back into DNA by a cellular reverse transcription (RT) process (also likely to be error-prone) we have called "RTmediated long DNA conversion" (RT-LDC). Finally we suggest that all these activities and others can be envisaged as being brought physically under the umbrella of special sites in the nucleus involved in transcription known as "Transcription Factories" (TF).
Recent innovations in microelectronics and advances in cryptography are driving the appearance of... more Recent innovations in microelectronics and advances in cryptography are driving the appearance of a new generation of smart cards with wider applications; this has important repercussions for our society in the coming years. Essentially, these breakthroughs include built-in microprocessors capable of generating cryptographic transactions (e.g.,Jelectronic blinded signatures, digital pseudonyms, and digital credentials), developments toward a single electronic card offering multi-access to services such as transport, telecommunications, health, financial, and entertainment (Universal Access Services), and incorporation of personal identification technologies such as voice, eye, or skin pattern recognition. For example, by using electronic representatives or cryptographic blinded signatures, a smart card can be used for multi transactions across different organizations and under different generated pseudonyms. These pseudonyms are capable of recognizing an individual unambiguously, while none of her records can be linked [1]. Moreover, tamper-proof electronic observers would make smart cards a very attractive technology for high-security based applications, such as those in the health care field. New trends in smart card technology offer excellent privacy and confidentiality safeguards. Therefore, smart cards constitute a promising technology for the health sector in Australia and other countries around the world in their pursuit of technology to support the delivery of quality care services. This paper addresses the main issues and the key design criteria which may be of strategic importance to the success of future smart card technology in the health care sector.
Evidence already exists that the activation-induced deaminase (AID)/APOBEC family constitutes a s... more Evidence already exists that the activation-induced deaminase (AID)/APOBEC family constitutes a set of differentially expressed enzymes capable of deaminating cytosines (C to U) in singlestranded DNA (ssDNA) and that they are potentially powerful mutagens. The mutagenic processes involved are believed to be activated in many nonlymphoid tissue typesdfor example, initiating some cancers and/or leading to further somatic mutagenesis. To investigate the extent that codon context might be important in influencing the likely location of TP53 mutations in breast cancer, the codon-bias patterns resulting from the ssDNA target specificities of cytidine deaminases of the AID/APOBEC family were analyzed. The data indicate that codon context strongly influences the likely location of mutations at motifs for AID/APOBEC1/APOBEC3G, and at WA sites. An unexpected finding is a highly significant preference for transitions of cytosine to occur at the first nucleotide position and for transitions of guanosine to occur at the second nucleotide position in the mutated codon (read 3 0 to 5 0). Thus, the mechanisms involved appear to be sensitive to codon reading frames and to have an intrinsic ability to differentiate between the cytosines on the nontranscribed strand and those on the transcribed strand in the context of an open "transcription bubble."
Mutation Research: Fundamental And Molecular Mechanisms Of Mutagenesis, Jul 1, 2018
Cytosine and adenosine deamination events (DNA, RNA substrates) account for most codon-context Ta... more Cytosine and adenosine deamination events (DNA, RNA substrates) account for most codon-context Targeted Somatic Mutation (TSM) patterns observed in immunoglobulin (Ig) somatic hypermutation (SHM), and in cancer exomes following Ig-SHM-like responses. TSM refers to the process of somatic mutagenesis involving deamination events that results on a dominant type of mutation (e.g., C-toT), and co-incident at a particular motif (e.g., WRC), and preferentially targeting the first, second or third nucleotide position within the mutated codon (e.g. MC1, MC2 or MC3, read 5-prime to 3-prime). It is now widely accepted that if left uncorrected, the accumulation of uncorrected TSMs involving the deaminases, may lead to a diagnosis of cancer or other degenerative disease. Our hypothesis is that many missense, nonsense and synonymous single nucleotide polymorphisms (SNPs) associated with clinically significant diseases may have arisen in the population by similar highly targeted deamination events. The OMIM database was searched for disease-associated SNPs on the X chromosome, and for all chromosomes. The nucleotide substitution patterns for disease-associated SNPs were analyzed by the TSM method to identify the likely deaminase source for C-to-U
This paper simply links the findings of Zheng, Lorenzo and Beal (2017) to our previous work on st... more This paper simply links the findings of Zheng, Lorenzo and Beal (2017) to our previous work on strand biased and codon-context mutation signatures in B lymphocytes (Ig SHM) and codon-contexted exomewide point mutation patterns in cancer genomes. We conclude that in vivo the A-to-I DNA editing component at RNA: DNA hybrids occurring in Transcription Bubbles, while important, is of far lower Ato-I editing efficiency than in dsRNA substrates (as shown in Zheng et al 2017). The extreme strand biased mutation patterns documented by us in vivo should be logically rationalized by the predicted sequential steps of the RNA/RT-based mechanism. Abbreviations used in this paper: Aag, alkyladenine DNA glycosylase; ADAR, Adenosine Deaminase that acts on RNA; ADAT, Adenosine Deaminase that acts on tRNA; AID, activation induced cytidine deaminase, a APOBEC family member, initiating via C-to-U lesions in ssDNA of class switch recombination (CSR) and somatic hypermutation (SHM) processes at somatically rearranged Ig V(D)J gene loci, and known to activate cytidine mutagenic deamination during transcription in other somatic tissues, particularly in cancer; APOBEC family, generic abbreviation for the deoxyribonucleic acid, or dCto-dU, deaminase family (APOBEC3 A, B, C, D, F, G, H) similar in DNA sequence to the "apolipoprotein B RNA editor" APOBEC1, and known to activate mutagenic cytidine deamination during transcription in somatic tissues, particularly in cancer; AP, an Abasic, or apurinic/apyrimidinic, site; APE, AP endonuclease; A-to-I, adenosine-toinosine RNA editing; BER, base excision repair; Deaminase, catalytic domain in ADAR and AID/APOBEC enzymes; DSB, double strand DNA breaks; Ig-SHM-like response, strand-biased somatic mutation patterns similar to that observed in Ig SHM; MMR, mismatch repair; Motif, 4 to 6 nucleotide (N) sequence defining specificity of deaminase targeting; MSH2-MSH6, MutSa heterodimer recognising mispaired bases in DNA duplex; N, any nucleotide; NTS, the non-transcribed, or "Top", strand; NGS, Next Generation Sequencing; Pol-h or DNA polymerase-h (eta); R, Adenosine (A) or Guanine (G) , purines; RNA Pol II, RNA Polymerase II; RT, reverse transcriptase; RT-Pol-h, reverse transcriptase activity displayed by Pol-h; S, strong base pair involving Cytosine (C) or Guanine (G); SHM, somatic hypermutation; T, Thymine; TS, the transcribed, or "Bottom", strand, in context of a Transcription Bubble; TSM, targeted somatic mutations : the process of targeting C and A nucleotides for deamination in actively transcribed genes that results in a dominant type of mutation caused by a DBD or Inf-DBD at a particular codon position; TSRT, target site reverse transcription; U, uracil; UNG, uracyl DNA glycosylase involved in BER at dU sites in DNA resulting in either an Abasic site (AP) or APE-mediated ssDNA nicks (above); UTR, untranslated regions in the upstream (5') and downstream (3') regulatory regions of protein coding genes; V(D)J, generic symbol for a rearranged immunoglobulin (or T cell receptor, TCR) variable region gene in the
Targeted Somatic Mutation (TSM) analysis for disease-associated SNPs on all genes that are: I X C... more Targeted Somatic Mutation (TSM) analysis for disease-associated SNPs on all genes that are: I X Chromosome OMIM SNPs; and, II All chromosome OMIM SNPs in Clinvar database.
This paper reports the results of our initial analysis of APOBEC and ADAR deaminase-mediated muta... more This paper reports the results of our initial analysis of APOBEC and ADAR deaminase-mediated mutation signature patterns in complete COVID-19 genomes from informative locations and times in China, USA and Spain in the 2019-2020 pandemic. We have identified a unique set of 'new' putative coordinated Riboswitches in COVID-19 genomes not previously identified, and likely generating variants of the known common strain Haplotypes now in circulation. The results reveal that COVID-19 diversifies using switching of RNA Haplotypes with minimal alteration to protein structure (the normal targets for B and T cells in conventional vaccine development). The deaminase-driven RNA Haplotypes are most likely aligned with RNA secondary structures as several studies already highlight how Riboswitches alter he ability of RNA to fold into intricate three-dimensional structures allowing them to execute their diverse cellular functions. The same functional outcomes are expected for viruses, partic...
In hepatocellular cancer (HCC) there is an over expression of the RNA editing enzyme ADAR1. Furth... more In hepatocellular cancer (HCC) there is an over expression of the RNA editing enzyme ADAR1. Further, the prominent genomic somatic mutation signature in HCC is almost exclusively focused on mutations at A:T base pairs where A-to-G mutations far exceed T-to-C mutations (when read on the non-transcribed strand). A clear mechanism for this extreme transcriptional strand biased mutation signature, putatively associated with over expression of ADAR1 deaminase, is yet to be explicitly demonstrated. The standard description of this strand bias has been nominally called “Transcription Coupled Damage” (TCD) to distinguish it from more conventional “Transcription Coupled Repair” (TCR). We show that the TCD description does not satisfy all features of the molecular evidence. The conventional view is that ADAR1 is thought to target adenosines at WA-sites for editing to inosine (I) in double stranded RNA stem-loop structures in transcripts. Here we show that the totality of the molecular and cel...
This chapter addresses the molecular mechanism and source of the numerous mutagenic changes that ... more This chapter addresses the molecular mechanism and source of the numerous mutagenic changes that genomes, particularly mammalian genomes, can undergo during normal development and in diseased states. The central role of pathogen and other disease-inducing innate immunity via the action of the cytosine (AID/APOBEC) and adenosine (ADAR) deaminases is reviewed in some depth. The general and universal nature of deaminase-mediated mutagenesis is an important key to understanding cosmic genetic evolutionary processes.
Cytosine and adenosine deamination events (DNA, RNA substrates) account for most codon-context Ta... more Cytosine and adenosine deamination events (DNA, RNA substrates) account for most codon-context Targeted Somatic Mutation (TSM) patterns observed in immunoglobulin (Ig) somatic hypermutation (SHM), and in cancer exomes following Ig-SHM-like responses. TSM refers to the process of somatic mutagenesis involving deamination events that results on a dominant type of mutation (e.g., C-to-T), and co-incident at a particular motif (e.g., WRC), and preferentially targeting the first, second or third nucleotide position within the mutated codon (e.g. MC1, MC2 or MC3, read 5-prime to 3-prime). It is now widely accepted that if left uncorrected, the accumulation of uncorrected TSMs involving the deaminases, may lead to a diagnosis of cancer or other degenerative disease. Our hypothesis is that many missense, nonsense and synonymous single nucleotide polymorphisms (SNPs) associated with clinically significant diseases may have arisen in the population by similar highly targeted deamination event...
Evidence already exists that the activation-induced cytidine deaminase (AID/APOBEC) and the adeno... more Evidence already exists that the activation-induced cytidine deaminase (AID/APOBEC) and the adenosine deaminase (ADAR) families of enzymes are implicated as powerful mutagens in oncogenic processes in many somatic tissues. Each deaminase is identified by the DNA or RNA nucleotide sequence ("motif") surrounding the nucleotide targeted for deamination. The primary objective of this study is to develop an in silico approach to identify nucleotide sequence changes of the target motifs of key deaminases during oncogenesis. If successful, a secondary objective is to investigate if such changes are associated with disease progression indicators that include disease stage and progression-free survival time. Using a discovery cohort of 194 high-grade serous ovarian adenocarcinoma (HGS-OvCa) exomes, the results confirm the ability of the novel in silico approach used to identify changes in the preferred target motifs for AID, APOBEC3G, APOBEC3B, and ADAR1 during oncogenesis. Using t...
There are strong indications that now is an appropriate time for introducing a health insurance s... more There are strong indications that now is an appropriate time for introducing a health insurance smart card in Australia. Reviews of health implementations in Europe to date verify that a critical mass of projects exists and that technological requirements have been satisfied. Costs of card readers have declined to a desirable level, the security features of smart card may be employed to overcome confidentiality and privacy concerns, and opportunities exist to improve the efficiency and cost effectiveness of health care delivery and administration on a national level. However for a successful health insurance smart card system to be implemented in Australia, unique socio-cultural and economic contexts must be incorporated into the systems design. These include implementation on a national scale to ensure coherency, compatibility, and interoperability between insurance and healthcare providers; modifying existing health information systems to migrate to the smart card network; convincing medical practitioners to adopt guidelines on electronic medical records; and ensuring privacy and confidentiality of patient records through a mixture of legislative, administrative, and technical controls. When these criteria are satisfied in the smart card insurance design, the opportunity for this technology to provide customised benefits, improved efficiency, and a speedier delivery of services in the current socioeconomic climate will be realised.
This thesis breaks new ground by providing the first detailed study of smart card innovation duri... more This thesis breaks new ground by providing the first detailed study of smart card innovation during its first twenty years (1974-1996). The overall aim is to apply sociotechnical principles to further our understanding of the innovation process as it relates to smart card technology. By using a sociotechnical framework, this study also seeks to illustrate the limitations of conventional innovation
Evidence already exists that the activation-induced deaminase (AID)/APOBEC family constitutes a s... more Evidence already exists that the activation-induced deaminase (AID)/APOBEC family constitutes a set of differentially expressed enzymes capable of deaminating cytosines (C to U) in single-stranded DNA (ssDNA) and that they are potentially powerful mutagens. The mutagenic processes involved are believed to be activated in many nonlymphoid tissue types-for example, initiating some cancers and/or leading to further somatic mutagenesis. To investigate the extent that codon context might be important in influencing the likely location of TP53 mutations in breast cancer, the codon-bias patterns resulting from the ssDNA target specificities of cytidine deaminases of the AID/APOBEC family were analyzed. The data indicate that codon context strongly influences the likely location of mutations at motifs for AID/APOBEC1/APOBEC3G, and at WA sites. An unexpected finding is a highly significant preference for transitions of cytosine to occur at the first nucleotide position and for transitions of...
Activation-induced cytidine deaminase (AID) initiates Phase I somatic hypermutation (SHM) of anti... more Activation-induced cytidine deaminase (AID) initiates Phase I somatic hypermutation (SHM) of antibody genes by deaminating deoxy-cytosine to deoxy-uracil (C-to-U). These lesions trigger Phase II, a poorly understood process of error-prone repair targeting AT pairs by DNA polymerase (Pol). Since Pol is also a reverse transcriptase, Phase II could involve copying off RNA as well as DNA templates. We explore this idea further since in an RNAbased pathway it is conceivable that adenosine-to-inosine (A-to-I) RNA editing causes Ato-G transitions since I like G pairs with C. Adenosine deaminases (ADARs) are known to preferentially edit A nucleotides that are preceded by an A or U (W) in double-stranded RNA substrates. On this assumption and using a theoretical bioinformatics approach we show that a significant and specific correlation (P < 0.002) exists between the frequency of WA-to-WG mutations and the number of mRNA hairpins that could potentially form at the mutation site. This implies roles for both RNA editing and reverse transcription during SHM in vivo and suggests definitive genetic experiments targeting the appropriate ADAR1 isoform (␥INF-ADAR1) and/or Ig pre-mRNA templates.
... systems design: an Australian case study JOAN COOPER, NILAY GENCTURK and ROBYN A. LINDLEY Dep... more ... systems design: an Australian case study JOAN COOPER, NILAY GENCTURK and ROBYN A. LINDLEY Department of Information and Communication Technology (IACT), The University of Wollongong, NorthŪelds Avenue, Wollongong, 2500 Australia Abstract. ...
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