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IP-MS/MS analysis of UV induced RPII215 interactome

ABSTRACT: Drosophila Melanogaster has been extensively used as a model system to study ionizing radiation and chemical induced mutagenesis, double strand break repair and recombination. However, there are only limited studies on nucleotide excision repair in this important model organism. In this study, we immunopreciptated DNA-directed RNA polymerase II (RPII215) complex from untreated and UV iradiated drosophila S2 cells and identified the protein that interact with it by mass spectrometry.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Drosophila Melanogaster (fruit Fly)

SUBMITTER: Li Wang

LAB HEAD: Aziz Sancar

PROVIDER: PXD028924 | Pride | 2022-04-04

REPOSITORIES: Pride

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			Action	DRS
	200225_AS_-UV_1_11_1.raw	Raw
	200225_AS_-UV_1_11_2.raw	Raw
	200225_AS_-UV_1_12_1.raw	Raw
	200225_AS_-UV_1_12_2.raw	Raw
	200225_AS_-UV_1_13_1.raw	Raw

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Publications

CSB-independent, XPC-dependent transcription-coupled repair in Drosophila.

Deger Nazli N Cao Xuemei X Selby Christopher P CP Gulec Saygin S Kawara Hiroaki H Dewey Evan B EB Wang Li L Yang Yanyan Y Archibald Sierra S Selcuk Berkay B Adebali Ogun O Sekelsky Jeff J Sancar Aziz A Liu Zhenxing Z

Proceedings of the National Academy of Sciences of the United States of America 20220301 9

Drosophila melanogaster has been extensively used as a model system to study ionizing radiation and chemical-induced mutagenesis, double-strand break repair, and recombination. However, there are only limited studies on nucleotide excision repair in this important model organism. An early study reported that Drosophila lacks the transcription-coupled repair (TCR) form of nucleotide excision repair. This conclusion was seemingly supported by the Drosophila genome sequencing p ...[more]

PMID: 35217627

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Modulation of UV damage formation and repair in nucleosomes in distinct chromatin states in Drosophila

Project description:Here, we describe a new genome-wide map of UV-induced cyclobutane pyrimidine dimers (CPDs) in Drosophila S2 cells and a naked DNA control using CPD-seq. We used this data to analyze CPD formation in nucleosomes and different chromatin states across the Drosophila genome. We analyzed CPD formation alongside existing excision repair-sequencing (XR-seq) data to compare CPD damage and repair rates in five distinct chromatin types in Drosophila. This analysis revealed that CPD repair varied in different chromatin domains, while CPD formation was largely unaffected. Moreover, we observed distinct patterns of repair activity in nucleosomes in different chromatin types.

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Combined transcriptome and proteome profiling indicate cell-type-specific functions of Drosophila garland and pericardial nephrocytes

Project description:Drosophila nephrocytes are specialised cells that share critical functional, morphological, and molecular features with podocytes of the mammalian kidney, including the slit diaphragm, which is a functional filtration barrier. Accordingly, nephrocytes are considered highly suitable invertebrate models for glomerular disease in humans. In this study, we established a method to individually isolate the garland and pericardial nephrocytes from Drosophila of different developmental stages. This allowed us to collect nephrocyte samples that were uncontaminated by other cell types. We used a combination of mass spectrometry-based proteomics and RNA-Seq-based transcriptomics to characterise the proteome and transcriptome of the respective cells in an integrated manner. In addition to identifying a set of factors that were expressed exclusively in nephrocytes, we observed characteristic changes in the cellular proteome and transcriptome as a consequence of ageing. Furthermore, functional enrichment analyses suggested that larval and adult nephrocytes fulfil distinct physiological functions, which was also indicated for garland and pericardial nephrocytes in adult animals. In addition, the latter cells were characterised by transcriptomic and proteomic profiles that were suggestive of a cell-type-specific energy metabolism. Finally, both types of nephrocytes displayed transcriptomic signatures that indicated elevated immune signalling. The data generated in this study may represent a valuable basis for a more specific application of the Drosophila model system in the analysis of renal cell function in health and disease.

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Multigenerational effect of heat stress on the Drosophila melanogaster sperm proteome

Project description:The notion that genes are the sole units of heredity and that a barrier exists between soma and germline has been a major hurdle in elucidating the heritability of traits that were observed to follow a non-Mendelian inheritance pattern. It was only after the conception of “epigenetics” by C. H. Waddington that the effect of parental environment on subsequent generations via non-DNA sequence-based mechanisms, such as DNA methylation, chromatin modifications, non-coding RNAs and proteins, could be established in various organisms, now referred to as multigenerational epigenetic inheritance. Despite the growing body of evidence, the male gamete-derived epigenetic factors that mediate the transmission of such phenotypes are seldom explored, particularly in the model organism Drosophila melanogaster. Using the heat stress-induced multigenerational epigenetic inheritance paradigm in a widely used position-effect variegation line of Drosophila, named white-mottled, we have dissected the effect of heat stress on the sperm proteome in the current study. We demonstrate that multiple successive generations of heat stress at the early embryonic stage results in a significant downregulation of proteins associated with translation, chromatin organization, microtubule-based processes, and generation of metabolites and energy in the Drosophila sperms. Based on our findings, we propose chromatin-based epigenetic mechanisms, a well-established mechanism for environmentally induced multigenerational effects, as a plausible way of transmitting heat stress memory via the male germ line in subsequent generations. Moreover, we demonstrate the effect of multiple generations of heat stress on the reproductive fitness of Drosophila, shedding light on the adaptive or maladaptive potential of heat stress-induced multigenerational phenotypes.

2024-05-10 | PXD037488 | Pride

Neprilysins regulate muscle contraction and heart function via cleavage of SERCA-inhibitory micropeptides

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2022-06-01 | PXD027738 | Pride

CPD-seq mapping of UV damage formation and repair in yeast

Project description:UV-induced DNA lesions are an important contributor to mutagenesis and cancer, but it is not fully understood how the chromosomal landscape influences UV lesion formation and repair. We have used a novel high-throughput sequencing method to precisely map UV-induced cyclobutane pyrimidine dimers (CPDs) at nucleotide resolution throughout the yeast genome. Analysis of CPD formation reveals that nucleosomal DNA having an inward rotational setting is protected from CPD lesions. In strongly positioned nucleosomes, this nucleosome 'photofootprint' overrides intrinsic dipyrimidine sequence preferences for CPD formation. CPD formation is also inhibited by DNA-bound transcription factors, in effect protecting important DNA elements from UV damage. Analysis of CPD repair revealed a clear signature of efficient transcription-coupled nucleotide excision repair. Repair was less efficient at translational positions near a nucleosome dyad and at heterochromatic regions in the yeast genome. These findings define the roles of nucleosomes and transcription factors in UV damage formation and repair. UV mapping data was analyzed for yeast cells irradiated with 125J/m2 and allowed to repair for 0hr (2 samples), 20 minutes, 1 hour, or 2 hours. Data is also included for naked DNA irradiated with UV 60 or 90 J/m2

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Genome-wide analysis of human global and transcription-coupled excision repair of UV damage at single-nucleotide resolution

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