Biology subjects

Vu, H.

Publications and source records attributed to Vu, H..

5 recordsLinked to original sources

bioRxiv2022

In-vivo characterization of glutamine metabolism identifies therapeutic targets in clear cell renal cell carcinoma

Targeting metabolic vulnerabilities has been proposed as a therapeutic strategy in renal cell carcinoma (RCC). Here, we analyzed metabolism in patient-derived xenografts (tumorgrafts) from diverse forms of RCC. Tumorgrafts from VHL-mutant clear cell RCC (ccRCC) retained metabolic features of human ccRCC and engage in oxidative and reductive glutamine metabolism. We used several approaches to suppress glutamine metabolism and test the effect on tumor growth. Genetic silencing of isocitrate dehydrogenase-1 or -2 impaired reductive labeling of TCA cycle intermediates and suppressed tumor growth. Glutaminase inhibition resulted in modest growth suppression and variable effects on glutamine metabolism in vivo. Infusions with [amide-15N]glutamine revealed persistent amidotransferase activity during glutaminase inhibition, and blocking these activities with the amidotransferase inhibitor JHU-083 also reduced tumor growth. We conclude that ccRCC tumorgrafts catabolize glutamine via multiple pathways, perhaps explaining why it has been challenging to achieve therapeutic responses in patients by inhibiting glutaminase. TeaserGlutamine fuels the TCA cycle and amidotransferase pathways in clear cell renal cell carcinoma.

cancer biology↗

bioRxiv2022

ERK2-topoisomerase II regulatory axis is important for gene activation in immediate early genes

The transcription of stress-inducible genes requires synchronized and robust activation, which is critical for organismal survival and homeostasis. The function of the mitogen-activated protein kinase (MAPK) signaling pathway is required for the activation of immediate early genes (IEGs), including EGR1 and FOS, for cell growth and proliferation1-3. In addition, recent studies have identified topoisomerase II (TOP2) as one of the important regulators of the transcriptional activation of IEGs4-6. However, the mechanism underlying transcriptional regulation involving TOP2 in IEG activation has remained unknown. Here, we demonstrate that ERK2, but not ERK1, is important for IEG transcriptional activation and report a critical ELK1 binding sequence for ERK2 function at the EGR1 gene. Our data indicate that both ERK1 and ERK2 extensively phosphorylate the C-terminal domain of TOP2B at mutual and distinctive residues. Although both ERK1 and ERK2 enhance the catalytic rate of TOP2B required to relax positive DNA supercoiling, ERK1 can relax the DNA by itself and produces a semi-relaxed DNA, which is apparently resistant to TOP2B catalysis. Inhibition of ERK2 kinase activity or ERK2 knock-down interferes with transcription and deregulates TOP2B in IEGs. Furthermore, we obtained the first cryo-EM structure of the human cell-purified TOP2B and etoposide together with the EGR1 transcriptional start site (50 nt; -30 to +20) that has the strongest affinity to TOP2B within -423 to +332. The structure elucidated in our studies showed TOP2B-mediated breakage and dramatic bending of the double-stranded DNA, comparable to previously reported structures of TOP2. Our cell-based analyses showed transcriptional activation by etoposide and transcriptional inhibition by ICRF193 at EGR1 and FOS, suggesting that TOP2B-mediated DNA break to favor transcriptional activation. Taken together, this study suggests that activated ERK2 phosphorylates TOP2B to regulate TOP2-DNA interactions and favor transcriptional activation in IEGs. We propose that TOP2B association, catalysis, and dissociation on its substrate DNA are important processes for regulating transcription and that ERK2-mediated TOP2B phosphorylation may be key for the catalysis and dissociation steps.

molecular biology↗

bioRxiv2022

Structural and mechanistic basis of σ-dependent transcriptional pausing

In {sigma}-dependent transcriptional pausing, the transcription initiation factor {sigma}, translocating with RNA polymerase (RNAP), makes sequence-specific protein-DNA interactions with a promoter-like sequence element in the transcribed region, inducing pausing. It has been proposed that, in {sigma}-dependent pausing, the RNAP active center can access off-pathway "backtracked" states that are substrates for the transcript-cleavage factors of the Gre family, and on-pathway "scrunched" states that mediate pause escape. Here, using site-specific protein-DNA photocrosslinking to define positions of the RNAP trailing and leading edges and of {sigma} relative to DNA at the{lambda} PR promoter, we show directly that {sigma}-dependent pausing in the absence of GreB in vitro predominantly involves a state backtracked by 2-4 bp, and that {sigma}-dependent pausing in the presence of GreB in vitro and in vivo predominantly involves a state scrunched by 2-3 bp. Analogous experiments with a library of 47 ([~]16,000) transcribed-region sequences show that the state scrunched by 2-3 bp--and only that state--is associated with the consensus sequence, T-3N-2Y-1G+1, (where -1 corresponds to the position of the RNA 3 end), which is identical to the consensus for pausing in initial transcription, and which is related to the consensus for pausing in transcription elongation. Experiments with heteroduplex templates show that sequence information at position T-3 resides in the DNA nontemplate strand. A cryo-EM structure of a complex engaged in {sigma}-dependent pausing reveals positions of DNA scrunching on the DNA nontemplate and template strands and suggests that position T-3 of the consensus sequence exerts its effects by facilitating scrunching.

biochemistry↗

bioRxiv2021

Epigenetic predictors of maximum lifespan and other life history traits in mammals

Maximum lifespan of a species is the oldest that individuals can survive, reflecting the genetic limit of longevity in an ideal environment. Here we report methylation-based models that accurately predict maximum lifespan (r=0.89), gestational time (r=0.96), and age at sexual maturity (r=0.87), using cytosine methylation patterns collected from over 12,000 samples derived from 192 mammalian species. Our epigenetic maximum lifespan predictor corroborated the extended lifespan in growth hormone receptor knockout mice and rapamycin treated mice. Across dog breeds, epigenetic maximum lifespan correlates positively with breed lifespan but negatively with breed size. Lifespan-related cytosines are located in transcriptional regulatory regions, such as bivalent chromatin promoters and polycomb-repressed regions, which were hypomethylated in long-lived species. The epigenetic estimators of maximum lifespan and other life history traits will be useful for characterizing understudied species and for identifying interventions that extend lifespan.

evolutionary biology↗

bioRxiv2021

Standardized genome-wide function prediction enables comparative functional genomics in plants: a new application area for Gene Ontologies

BackgroundGenome-wide gene function annotations are useful for hypothesis generation and for prioritizing candidate genes potentially responsible for phenotypes of interest. We functionally annotated the genes of 18 crop plant genomes across 14 species using the GOMAP pipeline. ResultsBy comparison to existing GO annotation datasets, GOMAP-generated datasets cover more genes, contain more GO terms, and produce datasets similar in quality (based on precision and recall metrics using existing gold standards as the basis for comparison). From there, we sought to determine whether the datasets across multiple species could be used together to carry out comparative functional genomics analyses in plants. To test the idea and as a proof of concept, we created dendrograms of functional relatedness based on terms assigned for all 18 genomes. These dendrograms were compared to well-established species-level evolutionary phylogenies to determine whether trees derived were in agreement with known evolutionary relationships, which they largely are. Where discrepancies were observed, we determined branch support based on jack-knifing then removed individual annotation sets by genome to identify the annotation sets causing unexpected relationships. ConclusionsGOMAP-derived functional annotations used together across multiple species generally retain sufficient biological signal to recover known phylogenetic relationships based on genome-wide functional similarities, indicating that comparative functional genomics across species based on GO data hold promise for generating novel hypotheses about comparative gene function and traits.

bioinformatics↗