Papers by Gerald Schumann
Approximately 2% oftheDictyostelium discoideum genome consistsof multiple copiesof a retrotranspo... more Approximately 2% oftheDictyostelium discoideum genome consistsof multiple copiesof a retrotransposable element termed DRE(Dictyostelium Repetitive Element). Theseelements havealways been foundintegrated inaposition andorientation-specific manner50 4nucleotides upstream ofthecoding region oftRNAgenes(tDNAs). Anintact DREis5.7kb long. Itcarries anextensive coding region flanked by non-identical longterminal repeats (LTRs), composed ofthree distinct modules A,B andC.Theleft LTR proximal tothetRNAgenecontains oneorseveral Amodulesfollowed bya single B-module (AnB). By contrast, theright LTRiscomposedofaB-module followed byaC-module (BC). Approximately 50%ofthe DREelements inNC4derivatives ofD.discoideum are structurally different fromthe5.7kbDREdescribed above. Theycarry thefollowing alterations: a)a3.1kb deletion inthecoding region; b)twosmall deletions of8and29nucleotides intheB-module oftheright LTR;c)a72bpdeletion intheB-Cjunction; andd)three distinct point mutations within theA-mod...
Nature Communications, 2018
Mobile DNA, 2019
Human stem cells harbor significant potential for basic and clinical translational research as we... more Human stem cells harbor significant potential for basic and clinical translational research as well as regenerative medicine. Currently~3000 adult and~30 pluripotent stem cell-based, interventional clinical trials are ongoing worldwide, and numbers are increasing continuously. Although stem cells are promising cell sources to treat a wide range of human diseases, there are also concerns regarding potential risks associated with their clinical use, including genomic instability and tumorigenesis concerns. Thus, a deeper understanding of the factors and molecular mechanisms contributing to stem cell genome stability are a prerequisite to harnessing their therapeutic potential for degenerative diseases. Chemical and physical factors are known to influence the stability of stem cell genomes, together with random mutations and Copy Number Variants (CNVs) that accumulated in cultured human stem cells. Here we review the activity of endogenous transposable elements (TEs) in human multipotent and pluripotent stem cells, and the consequences of their mobility for genomic integrity and host gene expression. We describe transcriptional and post-transcriptional mechanisms antagonizing the spread of TEs in the human genome, and highlight those that are more prevalent in multipotent and pluripotent stem cells. Notably, TEs do not only represent a source of mutations/CNVs in genomes, but are also often harnessed as tools to engineer the stem cell genome; thus, we also describe and discuss the most widely applied transposon-based tools and highlight the most relevant areas of their biomedical applications in stem cells. Taken together, this review will contribute to the assessment of the risk that endogenous TE activity and the application of genetically engineered TEs constitute for the biosafety of stem cells to be used for substitutive and regenerative cell therapies.
Journal of Neurology, Neurosurgery & Psychiatry, 2018
Endogenous retrotransposon sequences constitute approximately 42% of the human genome, and mobili... more Endogenous retrotransposon sequences constitute approximately 42% of the human genome, and mobilisation of retrotransposons has resulted in rearrangements, duplications, deletions, novel transcripts and the introduction of new regulatory domains throughout the human genome. Both germline and somatic de novo retrotransposition events have been involved in a range of human diseases, and there is emerging evidence for the modulation of retrotransposon activity during the development of specific diseases. Particularly, there is unequivocal consensus that endogenous retrotransposition can occur in neuronal lineages. This review addresses our current knowledge of the different mechanisms through which retrotransposons might influence the development of and predisposition to amyotrophic lateral sclerosis.
Is the human early embryo unique in lacking an inner cell mass (ICM) and having parallel rather t... more Is the human early embryo unique in lacking an inner cell mass (ICM) and having parallel rather than step-wise development? Here we reanalyse single-cell transcriptomic data and stain human embryos in situ to reveal both classical step-wise development and the missing ICM, a transcriptomic homolog of macaque ICM, that differentiates to epiblast and primitive endoderm. This apparent classicism obscures numerous features that render our blastocyst phylogenetically distinct: unlike mice, human epiblast has hallmarks of self-renewal and we have abundant, previously unrecognized, blastocyst non-committed cells (NCCs), part of an apoptosis-mediated quality control/purging process. Comparative transcriptomics further reveals the transcriptomes of the pluripotent cells to be especially fast evolving, rendering all primate embryos unique. Rapid transcriptome turnover is in large part owing to endogenous retrovirus H (ERVH) activity, ERVH being associated with recent major gene expression gai...
Arteriosclerosis, thrombosis, and vascular biology, Jan 4, 2018
One source of endogenous reverse transcriptase (eRT) activity in nucleated cells is the LINE-1/L1... more One source of endogenous reverse transcriptase (eRT) activity in nucleated cells is the LINE-1/L1 (long interspersed nuclear element-1), a non-LTR retrotransposon that is implicated in the regulation of gene expression. Nevertheless, the presence and function of eRT activity and LINE-1 in human platelets, an anucleate cell, has not previously been determined. We demonstrate that human and murine platelets possess robust eRT activity and identify the source as being LINE-1 ribonucleoprotein particles. Inhibition of eRT in vitro in isolated platelets from healthy individuals or in people with HIV treated with RT inhibitors enhanced global protein synthesis and platelet activation. If HIV patients were treated with reverse transcriptase inhibitor, we found that platelets from these patients had increased basal activation. We next discovered that eRT activity in platelets controlled the generation of RNA-DNA hybrids, which serve as translational repressors. Inhibition of platelet eRT li...
Nucleic Acids Research, 2011
SINE-VNTR-Alu (SVA) elements are nonautonomous, hominid-specific non-LTR retrotransposons and dis... more SINE-VNTR-Alu (SVA) elements are nonautonomous, hominid-specific non-LTR retrotransposons and distinguished by their organization as composite mobile elements. They represent the evolutionarily youngest, currently active family of human non-LTR retrotransposons, and sporadically generate disease-causing insertions. Since preexisting, genomic SVA sequences are characterized by structural hallmarks of Long Interspersed Elements 1 (LINE-1, L1)-mediated retrotransposition, it has been hypothesized for several years that SVA elements are mobilized by the L1 protein machinery in trans. To test this hypothesis, we developed an SVA retrotransposition reporter assay in cell culture using three different human-specific SVA reporter elements. We demonstrate that SVA elements are mobilized in HeLa cells only in the presence of both L1-encoded proteins, ORF1p and ORF2p. SVA trans-mobilization rates exceeded pseudogene formation frequencies by 12-to 300-fold in HeLa-HA cells, indicating that SVA elements represent a preferred substrate for L1 proteins. Acquisition of an AluSp element increased the trans-mobilization frequency of the SVA reporter element by $25-fold. Deletion of (CC CTCT) n repeats and Alu-like region of a canonical SVA reporter element caused significant attenuation of the SVA trans-mobilization rate. SVA de novo insertions were predominantly full-length, occurred preferentially in G+C-rich regions, and displayed all features of L1-mediated retrotransposition which are also observed in preexisting genomic SVA insertions.
Trends in Evolutionary Biology
The SVA family of hominid-specific non-LTR retrotransposon comprises the youngest group of transp... more The SVA family of hominid-specific non-LTR retrotransposon comprises the youngest group of transposable elements in the human genome. The propagation of the most ancient SVA subfamily took place about 13.5 million years ago, and the youngest SVA subfamily appeared in the human genome after the human/chimpanzee divergence. Functional analysis of genes associated with SVA insertions demonstrated their link to multiple ontological categories, with one of the major categories being attributed to brain function. Further analysis of this subset demonstrated that SVA elements expanded their presence in the human genome at different stages of hominoid evolution and were associated with progressively evolving behavioral features that indicate a potential impact of SVA propagation on the cognitive ability of a modern human. Our analysis suggests a potential role of SVAs in the evolution of human central nervous system and especially in the emergence of functional trends relevant to social and...
ABSTRACTAPOBEC3 deaminases (A3s) provide mammals with an anti-retroviral barrier by catalyzing dC... more ABSTRACTAPOBEC3 deaminases (A3s) provide mammals with an anti-retroviral barrier by catalyzing dC-to-dU deamination on viral ssDNA. Within primates, A3s have evolved diversely via gene duplications and fusions. Human APOBEC3C (hA3C) efficiently restricts the replication of viral infectivity factor (vif)-deficient Simian immunodeficiency virus (SIVΔvif), but for unknown reasons, it inhibits HIV-1Δvif weakly. In catarrhines (Old World monkeys and apes), the A3C loop 1 displays the conserved amino acid pair WE, while the corresponding consensus sequence in A3F and A3D is the largely divergent pair RK, which is also the inferred ancestral sequence for the last common ancestor of A3C|D|F in primates. Here, we report that modifying the WE residues in hA3C loop 1 to RK leads to stronger interactions with ssDNA substrate, facilitating catalytic function, which resulted in a drastic increase in both deamination activity and the ability to restrict HIV-1 and LINE-1 replication. Conversely, th...
Retrovirology, 2016
Question: Foamy viruses (FV), and in particular PFV, have emerged in recent years as attractive g... more Question: Foamy viruses (FV), and in particular PFV, have emerged in recent years as attractive gene therapy vector candidates. Since the lack of knowledge on molecular events in FV replication is a major hurdle for broader usage of foamy virus vectors, we aimed at elucidating PFV biology by investigating interactions of its capsid protein, Gag, with host cell components. Methodology and result: To this end, we identified members of the mammalian PLK family as PFV Gag interactants in a commercial yeasttwo-hybrid (Y2H) screen and validated these results in detailed Y2H experiments for PLK1-3. In the yeast system, the intact PLK kinase and substrate recognition motifs were required for interactions with PFV Gag, in which a unique S 224-T-P 226 motif served as a PLK binding determinant. PFV Gag mutants harbouring alanine substitutions of STP residues (iSTP) or phosphomimetic mutations of the T 225 (pmSTP) failed to interact with PLK1-3 in yeast. These findings were corroborated by colocalization studies of ectopically expressed, fluorescently tagged proteins in mammalian cells, where mCherry-tagged PFV Gag was able to recruit eGFP-tagged PLK1 and 2 to condensed mitotic chromatin in an STP motif-dependent manner. When characterizing PFV virions containing wild type or STP mutant Gag proteins, we observed that the mutations did not interfere with particle assembly, release or reverse transcription, but led to a 70 % titer reduction relative to wild type in single-round infection experiments. These replication defects became more prominent in the replication-competent PFV context. Therefore, the lack of Gag STP mutant interaction with PLK proteins upon viral entry into host cells was likely underlying this replication deficit. This hypothesis was strengthened by the finding that enzymatic PLK inhibition in host cells during transduction with wild type PFV mimicked the replication phenotype of PFV STP mutants. In addition to the overall reduced infectivity of the mutants, we also observed that the STP mutations in particle-associated Gag lead to differential sensitivity to integrase inhibition by dolutegravir and resulted in decreased integration efficiency. Conclusions: Taken together, our results demonstrate that PLK proteins influence PFV replication by virtue of their interaction with the Gag protein, ensuring timely and efficient transduction. O2 A novel entry/uncoating assay reveals the presence of at least two species of viral capsids during synchronized HIV-1 infection
PloS one, 2016
APOBEC4 (A4) is a member of the AID/APOBEC family of cytidine deaminases. In this study we found ... more APOBEC4 (A4) is a member of the AID/APOBEC family of cytidine deaminases. In this study we found a high mRNA expression of A4 in human testis. In contrast, there were only low levels of A4 mRNA detectable in 293T, HeLa, Jurkat or A3.01 cells. Ectopic expression of A4 in HeLa cells resulted in mostly cytoplasmic localization of the protein. To test whether A4 has antiviral activity similar to that of proteins of the APOBEC3 (A3) subfamily, A4 was co-expressed in 293T cells with wild type HIV-1 and HIV-1 luciferase reporter viruses. We found that A4 did not inhibit the replication of HIV-1 but instead enhanced the production of HIV-1 in a dose-dependent manner and seemed to act on the viral LTR. A4 did not show detectable cytidine deamination activity in vitro and weakly interacted with single-stranded DNA. The presence of A4 in virus producer cells enhanced HIV-1 replication by transiently transfected A4 or stably expressed A4 in HIV-susceptible cells. APOBEC4 was capable of similarl...
Molecular and cellular biology, 1992
Different Dictyostelium discoideum strains contain between 2 and 200 copies of a retrotransposabl... more Different Dictyostelium discoideum strains contain between 2 and 200 copies of a retrotransposable element termed DRE (Dictyostelium repetitive element). From the analysis of more than 50 elements, it can be concluded that DRE elements always occur 50 +/- 3 nucleotides upstream of tRNA genes. All analyzed clones contain DRE in a constant orientation relative to the tRNA gene, implying orientation specificity as well as position specificity. DRE contains two open reading frames which are flanked by nonidentical terminal repeats. Long terminal repeats (LTRs) are composed of three distinct modules, called A, B, and C. The tRNA gene-proximal LTR is characterized by one or multiple A modules followed by a single B module (AnB). With respect to the distal LTR, two different subforms of DRE have been isolated. The majority of isolated clones contains a distal LTR composed of a B module followed by a C module (BC), whereas the distal LTR of the other subform contains a consecutive array of ...
Genome biology and evolution, Oct 15, 2016
LAVA ( L: INE- A: lu- V: NTR- A: lu-like) elements comprise a family of non-autonomous, composite... more LAVA ( L: INE- A: lu- V: NTR- A: lu-like) elements comprise a family of non-autonomous, composite, non-LTR retrotransposons specific to gibbons and may have played a role in the evolution of this lineage. A full-length LAVA element consists of portions of repeats found in most primate genomes: CT-rich, Alu-like, and VNTR regions from the SVA retrotransposon, and portions of the AluSz and L1ME5 elements. To evaluate whether the gibbon genome currently harbors functional LAVA elements capable of mobilization by the endogenous LINE-1 (L1) protein machinery and which LAVA components are important for retrotransposition, we established a trans-mobilization assay in HeLa cells. Specifically, we tested if a full-length member of the older LAVA subfamily C that was isolated from the gibbon genome and named LAVAC, or its components, can be mobilized in the presence of the human L1 protein machinery. We show that L1 proteins mobilize the LAVAC element at frequencies exceeding processed pseudo...
Methods in molecular biology (Clifton, N.J.), 2016
Mammalian genomes harbor autonomous retrotransposons coding for the proteins required for their o... more Mammalian genomes harbor autonomous retrotransposons coding for the proteins required for their own mobilization, and nonautonomous retrotransposons, such as the human SVA element, which are transcribed but do not have any coding capacity. Mobilization of nonautonomous retrotransposons depends on the recruitment of the protein machinery encoded by autonomous retrotransposons. Here, we summarize the experimental details of SVA trans-mobilization assays which address multiple questions regarding the biology of both nonautonomous SVA elements and autonomous LINE-1 (L1) retrotransposons. The assay evaluates if and to what extent a noncoding SVA element is mobilized in trans by the L1-encoded protein machinery, the structural organization of the resulting marked de novo insertions, if they mimic endogenous SVA insertions and what the roles of individual domains of the nonautonomous retrotransposon for SVA mobilization are. Furthermore, the highly sensitive trans-mobilization assay can be...
Oncotarget, Jan 26, 2015
LINE-1 (L1) retrotransposons are a source of endogenous reverse transcriptase (RT) activity, whic... more LINE-1 (L1) retrotransposons are a source of endogenous reverse transcriptase (RT) activity, which is expressed as part of the L1-encoded ORF2 protein (L1-ORF2p). L1 elements are highly expressed in many cancer types, while being silenced in most differentiated somatic tissues. We previously found that RT inhibition reduces cell proliferation and promotes differentiation in neoplastic cells, indicating that high endogenous RT activity promotes cancer growth. Here we investigate the expression of L1-ORF2p in several human types of cancer.We have developed a highly specific monoclonal antibody (mAb chA1-L1) to study ORF2p expression and localization in human cancer cells and tissues.We uncover new evidence for high levels of L1-ORF2p in transformed cell lines and staged epithelial cancer tissues (colon, prostate, lung and breast) while no or only basal ORF2p expression was detected in non-transformed cells. An in-depth analysis of colon and prostate tissues shows ORF2p expression in p...
Journal of Virology
Capsid-targeted viral inactivation is a novel protein-based strategy for the treatment of viral i... more Capsid-targeted viral inactivation is a novel protein-based strategy for the treatment of viral infections. Virus particles are inactivated by targeting toxic fusion proteins to virions, where they destroy viral components from within. We have fused Staphylococcus nuclease (SN) to the C-terminal end of Moloney murine leukemia virus Gag and demonstrated that expression of this fusion protein in chronically infected chicken embryo fibroblasts resulted in its incorporation into virions and subsequent inactivation of the virus particles by degradation of viral RNA. Release of particles incorporating Gag-SN fusion proteins into the extracellular milieu activates the nuclease and results in destruction of the virion from within. By comparing the effects of incorporated SN and SN*, an enzymatically inactive missense mutant form of SN, on the infectivity of virus particles, we have clearly demonstrated that nucleolytic activity is the antiviral mechanism. Expression of Gag-SN fusion protein...
Cytogenetic and Genome Research, 2005
Non-long terminal repeat (Non-LTR) retrotransposons represent a diverse and widely distributed gr... more Non-long terminal repeat (Non-LTR) retrotransposons represent a diverse and widely distributed group of transposable elements and an almost ubiquitous component of eukaryotic genomes that has a major impact on evolution. Their copy number can range from a few to several million and they often make up a significant fraction of the genomes. The members of the dominating subtype of non-LTR retrotransposons code for an endonuclease with homology to apurinic/apyrimidinic endonucleases (APE), and are thus termed APE-type non-LTR retrotransposons. In the last decade both the number of identified non-LTR retrotransposons and our knowledge of biology and evolution of APE-type non-LTR retrotransposons has increased tremendously.
Archives of Virology, 1998
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Papers by Gerald Schumann