Biology subjects

Landsman, D.

Publications and source records attributed to Landsman, D..

10 recordsLinked to original sources

bioRxiv2024

Cancer Histone Mutations Impact Binding and DNA Repair Processes, Leading to Increased Mutagenesis

Histones are key epigenetic factors for regulating the accessibility and compaction of eukaryotic genomes, affecting the replication, repair, and expression of DNA. Recent studies have demonstrated that histone missense mutations can perturb normal histone function, promoting the development of phenotypically distinguishable cancers. However, most histone mutations observed in cancer patients remain enigmatic in their potential to promote cancer development. To assess the oncogenic potential of histone missense mutations, we have gathered whole-exome sequencing data for the tumors of over 12,000 patients. Overall, histone mutations occurred in about 16% of cancer patients, although specific cancer types showed substantially higher rates. Using a combination of genomic, structural, and biophysical analyses, we found several predominant modes of action, where cancer missense mutations in histones affected acidic patches and protein binding interfaces in a cancer-specific manner and targeted interaction sites with specific DNA repair proteins. Consistent with this finding, we observed a high tumour mutational burden in patients with histone mutations affecting interactions of DNA repair proteins. We also identified potential cancer driver mutations in several histone genes, including histone H4-a highly conserved histone without previously documented driver mutations.

bioinformatics↗

bioRxiv2023

Detection of new pioneer transcription factors as cell-type specific nucleosome binders

Wrapping of DNA into nucleosomes restricts accessibility to the DNA and may affect the recognition of binding motifs by transcription factors. A certain class of transcription factors, the pioneer transcription factors, can specifically recognize their DNA binding sites on nucleosomes, may initiate local chromatin opening and facilitate the binding of co-factors in a cell-type-specific manner. For the majority of human pioneer transcription factors, the locations of their binding sites, mechanisms of binding and regulation remain unknown. We have developed a computational method to predict the cell-type-specific ability of transcription factors to bind nucleosomes by integrating ChIP-seq, MNase-seq and DNase-seq data with details of nucleosome structure. We have demonstrated the ability of our approach in discriminating pioneer from canonical transcription factors and predicted new potential pioneer transcription factors in H1, K562, HepG2 and HeLa cell lines. Lastly, we systemically analyzed the interaction modes between various pioneer transcription factors and detected several clusters of distinctive binding sites on nucleosomal DNA.

bioinformatics↗

bioRxiv2022

De novo transcriptome assembly and the effect of foreign RNA contamination

Multiple next-generation-sequencing (NGS)-based studies are enabled by the availability of a reference genome of the target organism. Unfortunately, several organisms remain unannotated due to the cost and complexity of generating a complete (or close to complete) reference genome. These unannotated organisms, however, can also be studied if a de novo reference transcriptome is assembled from whole transcriptome sequencing experiments. This technology is cost effective and widely used but is susceptible to off-target RNA contamination. In this manuscript, we present GTax, a taxonomy structured database of genomic sequences that can be used with BLAST to detect and remove foreign contamination in RNA sequencing samples before assembly. In addition, we investigate the effect of foreign RNA contamination on a de novo transcriptome assembly of Solanum lycopersicum (tomato). Our study demonstrates that removing foreign contamination in sequencing samples reduces the number of assembled chimeric transcripts.

genomics↗

bioRxiv2021

DNA methylation cues in nucleosome geometry, stability, and unwrapping

Cytosine methylation at the 5-carbon position is an essential DNA epigenetic mark in many eukaryotic organisms. Although countless structural and functional studies of cytosine methylation have been reported in both prokaryotes and eukaryotes, our understanding of how it influences the nucleosome assembly, structure, and dynamics remains obscure. Here we investigated the effects of cytosine methylation at CpG sites on nucleosome dynamics and stability. By applying long molecular dynamics simulations (five microsecond long trajectories, 60 microseconds in total), we generated extensive atomic level conformational full nucleosome ensembles. Our results revealed that methylation induces pronounced changes in geometry for both linker and nucleosomal DNA, leading to a more curved, under-twisted DNA, shifting the population equilibrium of sugar-phosphate backbone geometry. These conformational changes are associated with a considerable enhancement of interactions between methylated DNA and the histone octamer, doubling the number of contacts at some key arginines. H2A and H3 tails play important roles in these interactions, especially for DNA methylated nucleosomes. This, in turn, prevents a spontaneous DNA unwrapping of 3-4 helical turns for the methylated nucleosome with truncated histone tails, otherwise observed in the unmethylated system on several microsecond time scale.

biophysics↗

bioRxiv2020

Binding of regulatory proteins to nucleosomes is modulated by dynamic histone tails

Despite histone tails critical roles in epigenetic regulation, little is known about mechanisms of how histone tails modulate the nucleosomal DNA solvent accessibility and recognition of nucleosomes by other macromolecules. Here we generate extensive atomic level conformational ensembles of histone tails in the context of the full human nucleosome, totaling 26 microseconds of molecular dynamics simulations. We explore the histone tail binding with the nucleosomal and linker DNA and observe rapid conformational transitions between bound and unbound states allowing us to estimate kinetic and thermodynamic properties of the histone tail-DNA interactions. Different histone types exhibit distinct, although conformationally heterogeneous, binding modes and each histone type occludes specific DNA regions from the solvent. Using a comprehensive set of experimental data on nucleosome structural complexes, we find that majority of the studied nucleosome-binding proteins and histone tails target mutually exclusive regions on nucleosomal or linker DNA around the super-helical locations {+/-}1, {+/-}2, and {+/-}7. This finding is explained within the generalized competitive binding and tail displacement models of partners recruitment to nucleosomes. Finally, we demonstrate the crosstalk between different histone post-translational modifications, where charge-altering modifications and mutations typically suppress tail-DNA interactions and enhance histone tail dynamics.

biophysics↗

bioRxiv2020

Altered Mediator dynamics during heat shock in budding yeast

The Mediator complex is central to transcription by RNA polymerase II (Pol II) in eukaryotes. In yeast, Mediator is recruited by activators via its tail module and then facilitates assembly of the pre-initiation complex (PIC), including Pol II, setting the stage for productive transcription. Mediator occupies proximal promoter regions only transiently prior to Pol II escape; interruption of the transcription cycle by inactivation or depletion of Kin28 inhibits Pol II escape and stabilizes Mediator occupancy at promoters. However, whether Mediator occupancy and dynamics differ for gene cohorts induced by stress or alternative growth conditions has not been examined on a genome-wide scale. Here we investigate Mediator occupancy following heat shock or CdCl2 induction, with or without depletion of Kin28. We find that Pol II occupancy exhibits similar dependence on Mediator under normal and heat shock conditions; however, Mediator occupancy does not increase upon Kin28 depletion at most genes active during heat shock, indicating altered dynamics. Furthermore, Mediator occupancy persists at genes repressed by heat shock or CdCl2 induction and exhibits peaks upstream of the proximal promoter whether or not Kin28 is depleted, suggesting that Mediator is recruited by activators but is unable to engage PIC components at these repressed targets. Finally, we show a reduced dependence on PIC components for Mediator occupancy at promoters after heat shock, further supporting an altered dynamics or stronger engagement with activators under these conditions.

molecular biology↗

bioRxiv2019

BAMscale: quantification of DNA sequencing peaks and generation of scaled coverage tracks

BAMscale is a one-step tool that processes DNA sequencing datasets from chromatin binding (ChIP-seq) and chromatin state changes (ATAC-seq, END-seq) experiments to DNA replication data (OK-seq, NS-seq and replication timing). The outputs include normalized peak scores in text format and scaled coverage tracks (BigWig) which are directly accessible to data visualization programs. BAMscale (available at https://github.com/ncbi/BAMscale) effectively processes large sequencing datasets (~100Gb size) in minutes, outperforming currently available tools.

bioinformatics↗

bioRxiv2019

Kin28 depletion increases association of TFIID subunits Taf1 and Taf4 with promoters in Saccharomyces cerevisiae

In eukaryotes, transcription of mRNA-encoding genes by RNA polymerase II (Pol II) begins with assembly of the pre-initiation complex (PIC), comprising Pol II and the general transcription factors. Although the pathway of PIC assembly is well established, the mechanism of assembly and the dynamics of PIC components are not fully understood. For example, only recently has it been shown in yeast that the Mediator complex, which assists in pre-initiation complex formation at promoters of essentially all genes transcribed by Pol II, normally occupies promoters only transiently. This was inferred from studies showing that inhibiting Pol II promoter escape by depleting or inactivating Kin28 resulted in increased promoter occupancy by Mediator, as measured by chromatin immunoprecipitation (ChIP). Here we show that two subunits of TFIID, Taf1 and Taf4, similarly show increased occupancy as measured by ChIP upon depletion or inactivation of Kin28. In contrast, TBP occupancy is unaffected by depletion of Kin28, thus revealing an uncoupling of Taf and TBP occupancy during the transcription cycle. Increased Taf1 occupancy upon Kin28 depletion is suppressed by depletion of TBP, while depletion of TBP in the presence of Kin28 has little effect on Taf1 occupancy. Taf1 occupancy relative to TBP is higher at TFIID-dominated promoters and promoters having consensus TATA elements than at SAGA-dominated promoters and promoters lacking consensus TATA elements, consistent with prior work, and the increase in Taf occupancy upon depletion of Kin28 is more pronounced at TFIID-dominated promoters. Our results support the suggestion, based on recent structural studies, that TFIID may not remain bound to gene promoters through the transcription initiation cycle.\n\nAuthor SummaryTranscription of mRNA-encoding genes by RNA polymerase II (Pol II) begins when the pre-initiation complex, a large complex comprising Pol II and several general transcription factors, including the TATA-binding protein (TBP)-containing TFIID complex, assembles at gene promoters. Although the major steps in the pathway of PIC assembly have been identified, the mechanism of assembly in vivo and the dynamics of PIC components are not fully understood. In this work we have used a yeast strain that is engineered to allow inhibition of promoter escape by Pol II by administration of a chemical, in order to \"freeze\" the assembled PIC and thus determine whether this condition increases the promoter occupancy of TBP and two TBP-associated factors (Tafs) that are components of TFIID. This approach was used recently to demonstrate that the Mediator complex, which facilitates PIC assembly, normally binds only transiently to gene promoters. We find that Tafs, like Mediator, show increased occupancy when Pol II promoter escape is inhibited, whereas TBP binding is constant. These results imply that binding of TBP and Tafs is uncoupled during the transcription cycle, and that Taf occupancy is at least partially interrupted upon Pol II promoter escape.

genetics↗

bioRxiv2017

Role of the pre-initiation complex in Mediator recruitment and dynamics

The Mediator complex functions in eukaryotic transcription by stimulating the cooperative assembly of a pre-initiation complex (PIC) and recruitment of RNA Polymerase II (Pol II) for gene activation. The core Mediator complex is organized into head, middle, and tail modules, and in budding yeast (Saccharomyces cerevisiae), Mediator recruitment has generally been ascribed to sequence-specific activators engaging the tail module triad of Med2-Med3-Med15 at upstream activating sequences (UASs). We show that med2{Delta} med3{Delta} med15{Delta} yeast are viable and that Mediator lacking Med2-Med3-Med15 is associated with active promoters genome-wide. To test whether Mediator might alternatively be recruited via interactions with the PIC, we examined Mediator association genome-wide after depleting PIC components. We found that depletion of Taf1, Rpb3, and TBP profoundly affected Mediator association at active gene promoters, with TBP being critical for transit of Mediator from UAS to promoter, while Pol II and Taf1 stabilize Mediator association at proximal promoters.

genetics↗

bioRxiv2017

The Mediator co-activator complex regulates Ty1 retromobility via alternative transcription start site selection

The Ty1 retrotransposons present in the genome of Saccharomyces cerevisiae belong to the large class of mobile genetic elements that replicate via an RNA intermediary and constitute a significant portion of most eukaryotic genomes. The retromobility of Ty1 is regulated by numerous host factors, including several subunits of the Mediator transcriptional co-activator complex. In spite of its known function in the nucleus, previous studies have implicated Mediator in the regulation of post-translational steps in Ty1 retromobility. To resolve this paradox, we systematically examined the effects of deleting nonessential Mediator subunits on the frequency of Ty1 retromobility and levels of retromobility intermediates. Our findings reveal that loss of distinct Mediator subunits alters Ty1 retromobility positively or negatively over a >10,000-fold range by regulating the ratio of an internal transcript, Ty1i, to the genomic Ty1 transcript. Ty1i RNA encodes a dominant negative inhibitor of Ty1 retromobility that blocks virus-like particle maturation and cDNA synthesis. These results resolve the conundrum of Mediator exerting sweeping control of Ty1 retromobility with only minor effects on the levels of Ty1 genomic RNA and the capsid protein, Gag. Since the majority of characterized intrinsic and extrinsic regulators of Ty1 retromobility alter a post-translational step(s), Mediator could play a central role in integrating signals that influence Ty1i expression to modulate retromobility.\n\nAuthor SummaryRetrotransposons are mobile genetic elements that copy their RNA genomes into DNA and insert the DNA copies into the host genome. These elements contribute to genome instability, control of host gene expression and adaptation to changing environments. Retrotransposons depend on numerous host factors for their own propagation and control. The retrovirus-like retrotransposon, Ty1, in the yeast Saccharomyces cerevisiae has been an invaluable model for retrotransposon research, and hundreds of host factors that regulate Ty1 retrotransposition have been identified. Non-essential subunits of the Mediator transcriptional co-activator complex have been identified as one set of host factors implicated in Ty1 regulation. Here, we report a systematic investigation of the effects of loss of these non-essential subunits of Mediator on Ty1 retrotransposition. Our findings reveal a heretofore unknown mechanism by which Mediator influences the balance between transcription from two promoters in Ty1 to modulate expression of an autoinhibitory transcript known as Ty1i RNA. Our results provide new insights into host control of retrotransposon activity via promoter choice and elucidate a novel mechanism by which the Mediator co-activator governs this choice.

molecular biology↗