Bioinformatics and Analytics Research Collaborative

Spt6-Spn1 interaction is required for RNA Polymerase II association and precise nucleosome positioning along transcribed genes

Tyler Interrante — Mon, 24 Mar 2025 13:59:36 +0000

Abstract

Spt6-Spn1 is an essential histone chaperone complex that associates with RNA Polymerase II (RNAPII) and reassembles nucleosomes during gene transcription. While the interaction between Spt6 and Spn1 is important for its histone deposition and transcription functions, a precise mechanistic understanding is still limited. Here, using temperature sensitive alleles of spt6 and spn1 that disrupt their interaction in yeast, we show that Spt6-Spn1 association is important for its stable interaction with the elongating RNAPII complex and nucleosomes. Using micrococcal nuclease (MNase)-based chromatin occupancy profiling, we further find that Spt6-Spn1 interaction is required to maintain a preferred nucleosome positioning at actively transcribed genes; in the absence of Spt6-Spn1 interaction, we observe a return to replication-dependent phasing. In addition to positioning defects, Spt6-Spn1 disrupting mutants also resulted in an overall shift of nucleosomes towards the 5’ end of genes that was correlated with decreased RNAPII levels. As loss of Spt6-Spn1 association results in cryptic transcription at a subset of genes, we examined these genes for their nucleosome profiles. These findings revealed that the chromatin organization at these loci is similar to other active genes, thus underscoring the critical role of DNA sequence in mediating cryptic transcription when nucleosome positioning is altered. Taken together, these findings reveal Spt6-Spn1 interaction is key to its association with elongating RNAPII and for its ability to precisely organize nucleosomes across transcription units.

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Early life exposure to vitamin D deficiency impairs molecular mechanisms that regulate liver cholesterol biosynthesis, energy metabolism, inflammation, and detoxification

Tyler Interrante — Fri, 10 May 2024 17:59:17 +0000

Introduction: Emerging data suggests liver disease may be initiated during development when there is high genome plasticity and the molecular pathways supporting liver function are being developed.

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CHD4 and SMYD1 repress common transcriptional programs in the developing heart

Tyler Interrante — Fri, 10 May 2024 17:57:40 +0000

Abstract

Regulation of chromatin states is essential for proper temporal and spatial gene expression. Chromatin states are modulated by remodeling complexes composed of components that have enzymatic activities. CHD4 is the catalytic core of the nucleosome remodeling and deacetylase (NuRD) complex, which represses gene transcription. However, it remains to be determined how CHD4, a ubiquitous enzyme that remodels chromatin structure, functions in cardiomyocytes to maintain heart development. In particular, whether other proteins besides the NuRD components interact with CHD4 in the heart is controversial. Using quantitative proteomics, we identified that CHD4 interacts with SMYD1, a striated muscle-restricted histone methyltransferase that is essential for cardiomyocyte differentiation and cardiac morphogenesis. Comprehensive transcriptomic and chromatin accessibility studies of Smyd1 and Chd4 null embryonic mouse hearts revealed that SMYD1 and CHD4 repress a group of common genes and pathways involved in glycolysis, response to hypoxia, and angiogenesis. Our study reveals a mechanism by which CHD4 functions during heart development, and a previously uncharacterized mechanism regarding how SMYD1 represses cardiac transcription in the developing heart.

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The tardigrade Hypsibius exemplaris dramatically upregulates DNA repair pathway genes in response to ionizing radiation

Tyler Interrante — Mon, 15 Apr 2024 16:22:25 +0000

Summary

Tardigrades can survive remarkable doses of ionizing radiation, up to about 1,000 times the lethal dose for humans. How they do so is incompletely understood. We found that the tardigrade Hypsibius exemplaris suffers DNA damage upon gamma irradiation, but the damage is repaired. We show that this species has a specific and robust response to ionizing radiation: irradiation induces a rapid upregulation of many DNA repair genes. This upregulation is unexpectedly extreme—making some DNA repair transcripts among the most abundant transcripts in the animal. By expressing tardigrade genes in bacteria, we validate that increased expression of some repair genes can suffice to increase radiation tolerance. We show that at least one such gene is important �� for tardigrade radiation tolerance. We hypothesize that the tardigrades’ ability to sense ionizing radiation and massively upregulate specific DNA repair pathway genes may represent an evolved solution for maintaining DNA integrity.

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Von Hippel Lindau tumor suppressor controls m6A-dependent gene expression in renal tumorigenesis

Tyler Interrante — Mon, 15 Apr 2024 16:20:43 +0000

Abstract

N6-Methyladenosine (m6A) is the most abundant posttranscriptional modification, and its contribution to cancer evolution has recently been appreciated. Renal cancer is the most common adult genitourinary cancer, approximately 85% of which is accounted for by the clear cell renal cell carcinoma (ccRCC) subtype characterized by VHL loss. However, it is unclear whether VHL loss in ccRCC affects m6A patterns. In this study, we demonstrate that VHL binds and promotes METTL3/METTL14 complex formation while VHL depletion suppresses m6A modification, which is distinctive from its canonical E3 ligase role. m6A RNA immunoprecipitation sequencing (RIP-Seq) coupled with RNA-Seq allows us to identify a selection of genes whose expression may be regulated by VHL-m6A signaling. Specifically, PIK3R3 is identified to be a critical gene whose mRNA stability is regulated by VHL in a m6A-dependent but HIF-independent manner. Functionally, PIK3R3 depletion promotes renal cancer cell growth and orthotopic tumor growth while its overexpression leads to decreased tumorigenesis. Mechanistically, the VHL-m6A–regulated PIK3R3 suppresses tumor growth by restraining PI3K/AKT activity. Taken together, we propose a mechanism by which VHL regulates m6A through modulation of METTL3/METTL14 complex formation, thereby promoting PIK3R3 mRNA stability and protein levels that are critical for regulating ccRCC tumorigenesis.

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Gastrulation-stage alcohol exposure induces similar rates of craniofacial malformations in male and female C57BL/6J mice

Tyler Interrante — Wed, 20 Dec 2023 15:22:32 +0000

Background

Prenatal alcohol exposure during gastrulation (embryonic day [E] 7 in mice, ~3rd week of human pregnancy) impairs eye, facial, and cortical development, recapitulating birth defects characteristic of Fetal Alcohol Syndrome (FAS). However, it is not known whether the prevalence or severity of craniofacial features associated with FAS is affected by biological sex.

Methods

The current study administered either alcohol (2.9 g/kg, two i.p. doses, 4 hr apart) or vehicle to pregnant C57BL/6J females on E7, prior to gonadal sex differentiation, and assessed fetal morphology at E17.

Results

Whereas sex did not affect fetal size in controls, alcohol-exposed females were smaller than both control females and alcohol-treated males. Alcohol exposure increased the incidence of eye defects to a similar degree in males and females. Together, these data suggest that females might be more sensitive to the general developmental effects of alcohol, but not effects specific to the craniofacies. Whole transcriptomic analysis of untreated E7 embryos found 214 differentially expressed genes in females vs. males, including those in pathways related to cilia and mitochondria, histone demethylase activity, and pluripotency.

Conclusion

Gastrulation-stage alcohol induces craniofacial malformations in male and female mouse fetuses at similar rates and severity, though growth deficits are more prevalent females. These findings support the investigation of biological sex as a contributing factor in prenatal alcohol studies.

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New BARC article: Gestational diabetes mellitus placentas exhibit epimutations at placental development genes

Paul Grant — Thu, 20 Oct 2022 18:57:15 +0000

Gestational diabetes mellitus (GDM) is a maternal metabolic disorder that perturbs placental development and increases the risk of offspring short- and long-term metabolic disorders. The mechanisms by which GDM impairs placental development remain poorly understood. Here, we defined the DNA methylome of GDM placentas and determined whether GDM perturbs methylation at genes important for placental development. We conducted an epigenome-wide association study of 42 placentas from pregnancies in the South African Soweto First 1000 days cohort (S1000). Using genome-wide bisulfite sequencing, we compared non-GDM placentas to GDM placentas with similar proportions from obese and non-obese mothers. Compared to non-GDM, GDM placentas exhibited a distinct methylation profile consisting of 12,210 differentially methylated CpGs (DMCs) that mapped to 3,875 genes. Epigenetically altered genes were enriched in Wnt and cadherin signalling pathways, both critical in placentation and embryogenesis. We also defined regional DNA methylation perturbation in GDM placentas at 11 placental development genes. These findings reveal extensive changes to the placental epigenome of GDM pregnancies and highlight perturbation enriched at important placental development genes. These molecular changes represent potential mechanisms for GDM-induced placental effects that may serve as candidate biomarkers for placental, maternal, and foetal health. Using a study design that used similar proportions of obese and non-obese mothers in our case and control pregnancies, we minimized the detection of changes due to obesity alone. Further work will be necessary to investigate the extent of the influence of obesity on these GDM-related placental epigenetic changes.

��ɴǰ��:��DNA methylation; Epigenetics; development; developmental programming; gestational diabetes mellitus; placenta.

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BARC Research Associate position open!

Paul Grant — Thu, 20 Oct 2022 18:56:01 +0000

�ճ�� Integrated Genomics Cores and Bioinformatic and Analytics Research Collaborative (BARC) is looking for a Research Associate/Analyst.

The analyst will provide bioinformatics and analytical support to a large cancer-focused project, and act as a consultant and analyst to projects in the �� research community for BARC and a precision medicine initiative (/ppmh/).

Of top priority BARC projects are COVID-19 genomics analyses, such as those on resistance mutation detection, host-pathogen interactions, and innate immune response to the virus in humans. Additionally, the analyst will be applying data science concepts and machine learning algorithms to electronic health record (EHR) data to solve clinical informatics problems for a precision medicine initiative. Other BARC projects will include DNA-Seq and RNA-Seq analyses, custom downstream analysis, and data visualization.

This position will spend 35% of its time providing bioinformatics and analytics support to cancer studies that use RNA-Seq data, 35% of its time consulting and providing bioinformatics support for COVID-19 studies and others from BARC, 15% of its time applying data science to projects on precision medicine and EHR data, and 15% of its time coordinating internships in the BARC. The Research Associate will be expected to provide practical mentorship and training to BARC Interns and support workshops on bioinformatics.

Familiarity with genomic data, electronic health record data and health informatics is strongly preferred; background in biology is helpful but not required. Must be able to work independently and as a part of a diverse team.

Skills:
– Advanced ability to code in a scripting language (Python or Perl), Pandas
– Proficiency with R or similar statistical language
–��SQL is a plus
– Experience working in a HPC��Ա��ǲԳ��Գ�
– Familiarity with current open-source bioinformatics tools for sequencing analysis
– Familiarity with electronic health record data a plus.

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New BARC article in NEJ: Enterovirus D68 in the Anterior Horn Cells of a Child with Acute Flaccid Myelitis

Paul Grant — Mon, 12 Sep 2022 16:57:11 +0000

To the Editor: Acute flaccid myelitis (AFM) is a
poliomyelitis-like paralyzing illness in children.
Since an outbreak of AFM in the United States in
2014, evidence has emerged suggesting that enterovirus D68 (EV-D68) causes AFM.1
The virus has
been detected in respiratory specimens obtained
from patients with AFM, but it has rarely been
detected in the cerebrospinal fluid. There have
been few autopsy studies of the disorder, and its
pathogenesis has been inferred mainly from in
vivo or in vitro models of EV-D68 infection or
historical comparisons with poliovirus infection

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Multivalent DNA and nucleosome acidic patch interactions specify VRK1 mitotic localization and activity

Paul Grant — Mon, 11 Apr 2022 13:52:24 +0000

A key role of chromatin kinases is to phosphorylate histone tails during mitosis to spatiotemporally regulate cell division. Vaccinia-related kinase 1 (VRK1) is a serine-threonine kinase that phosphorylates histone H3 threonine 3 (H3T3) along with other chromatin-based targets. While structural studies have defined how several classes of histone-modifying enzymes bind to and function on nucleosomes, the mechanism of chromatin engagement by kinases is largely unclear. Here, we paired cryo-electron microscopy with biochemical and cellular assays to demonstrate that VRK1 interacts with both linker DNA and the nucleosome acidic patch to phosphorylate H3T3. Acidic patch binding by VRK1 is mediated by an arginine-rich flexible C-terminal tail. Homozygous missense and nonsense mutations of this acidic patch recognition motif in VRK1 are causative in rare adult-onset distal spinal muscular atrophy. We show that these VRK1 mutations interfere with nucleosome acidic patch binding, leading to mislocalization of VRK1 during mitosis, thus providing a potential new molecular mechanism for pathogenesis.

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