Biology subjects

Ferrell, J.

Publications and source records attributed to Ferrell, J..

5 recordsLinked to original sources

bioRxiv2024

Mechanistic origins of temperature scaling in the early embryonic cell cycle

Temperature profoundly impacts organismal physiology and ecological dynamics, particularly affecting ectothermic species and making them especially vulnerable to climate changes. Although complex physiological processes usually involve dozens of enzymes, empirically it is found that the rates of these processes often obey the Arrhenius equation, which was originally proposed for individual chemical reactions. Here we have examined the temperature scaling of the early embryonic cell cycle, with the goal of understanding why the Arrhenius equation approximately holds and why it breaks down at temperature extremes. Using experimental data from Xenopus laevis, Xenopus tropicalis, and Danio rerio, plus published data from Caenorhabditis elegans, Caenorhabditis briggsae, and Drosophila melanogaster, we find that the apparent activation energies (Ea values) for the early embryonic cell cycle for diverse ectotherms are all similar, 75 {+/-} 7 kJ/mol (mean {+/-} std.dev., n = 6), which corresponds to a Q10 value at 20{degrees}C of 2.8 {+/-} 0.2 (mean {+/-} std.dev., n = 6). Using computational models, we find that the approximate Arrhenius scaling and the deviations from it at high and low temperatures can be accounted for by biphasic temperature scaling in critical individual components of the cell cycle oscillator circuit, by imbalances in the Ea values for different partially rate-determining enzymes, or by a combination of both. Experimental studies of cycling Xenopus extracts indicate that both of these mechanisms contribute to the general scaling of temperature, and in vitro studies of individual cell cycle regulators confirm that there is in fact a substantial imbalance in their Ea values. These findings provide mechanistic insights into the dynamic interplay between temperature and complex biochemical processes, and into why biological systems fail at extreme temperatures.

systems biology↗

bioRxiv2024

Long range regulation of transcription scales with genomic distance in a gene specific manner

While critical for tuning the timing and level of transcription, enhancer communication with distal promoters is not well understood. Here we bypass the need for sequence-specific transcription factors and recruit activators directly using CARGO-VPR, an approach for targeting dCas9-VPR using a multiplexed array of RNA guides. We show that this approach achieves effective activator recruitment to arbitrary genomic sites, even those inaccessible by single dCas9. We utilize CARGO-VPR across the Prdm8-Fgf5 locus in mESCs, where neither gene is expressed. We demonstrate that while activator recruitment to any tested region results in transcriptional induction of at least one gene, the expression level strongly depends on the genomic distance between the promoter and activator recruitment site. However, the expression-distance relationship for each gene scales distinctly in a manner not attributable to differences in 3D contact frequency, promoter DNA sequence or presence of the repressive chromatin marks at the locus.

molecular biology↗

bioRxiv2023

Deletion of smooth muscle O-GlcNAc transferase prevents development of atherosclerosis in western diet-fed hyperglycemic ApoE-/- mice in vivo

Accumulating evidence highlights protein O-GlcNAcylation as a putative pathogenic contributor of diabetic vascular complications. We previously reported that elevated protein O-GlcNAcylation correlates with increased atherosclerotic lesion formation and VSMC proliferation in response to hyperglycemia. However, the role of O-GlcNAc transferase (OGT), regulator of O-GlcNAc signaling, in evolution of diabetic atherosclerosis remains elusive. The goal of this study was to determine whether smooth muscle OGT (smOGT) plays a direct role in hyperglycemia-induced atherosclerotic lesion formation and SMC de-differentiation. Using tamoxifen-inducible Myh11-CreERT2 and OGTfl/fl mice, we generated smOGTWT and smOGTKO mice, with and without ApoE-null backgrounds. Following STZ-induced hyperglycemia, smOGTWT and smOGTKO mice were kept on standard laboratory diet for study duration. In a parallel study, smOGTWTApoE-/- and smOGTKOApoE-/- were kept on Western diet beginning 8-wks-age. Animals harvested at 14-16-wks-age were used for plasma and tissue collection. Loss of smOGT augmented SM contractile marker expression in aortic vessels of STZ-induced hyperglycemic smOGTKO mice. Consistently, smOGT deletion attenuated atherosclerotic lesion lipid burden (Oil red O), plaque area (H&E), leukocyte (CD45) and smooth muscle cell (ACTA2) abundance in Western diet-induced hyperglycemic smOGTKOApoE-/- mice. This was accompanied with increased SM contractile markers, and reduced inflammatory and proliferative marker expression. Further, smOGT deletion attenuated YY1 and SRF expression (transcriptional regulators of SM contractile genes) in hyperglycemic smOGTKOApoE-/- and smOGTKO mice. These data uncover an atheroprotective outcome of smOGT loss-of-function and suggest a direct regulatory role of OGT-mediated O-GlcNAcylation in VSMC de-differentiation in hyperglycemia.

pathology↗

bioRxiv2021

Stepwise Oxidations Play Key Roles in the Structural and Functional Regulations of DJ-1

DJ-1 is known to play neuroprotective roles by eliminating reactive oxygen species (ROS) as an antioxidant protein. However, the molecular mechanism of DJ-1 function has not been well elucidated. This study explored the structural and functional changes of DJ-1 in response to oxidative stress. We found that Cys46 is also reactive cysteine residue in DJ-1, which was identified employing an NPSB-B chemical probe that selectively reacts with redox sensitive cysteine sulfhydryl. Peroxidatic Cys46 readily formed an intra-disulfide bond with resolving Cys53, which was identified with nanoUPLC-ESI-q-TOF tandem mass spectrometry (MS/MS) employing DBond algorithm under the non-reducing condition. We also found that Cys46-Cys53 disulfide crosslinking affects the oxidative state of the third Cys106, which shows the crosstalk among three cysteine residues of DJ-1. Furthermore, we demonstrated that DJ-1 C46A mutant, not forming Cys46-Cys53 intra-disulfide bond, lost structural stability of DJ-1 employing hydrogen/deuterium exchange-mass spectrometry (HDX-MS) analysis. All three Cys mutants lost antioxidant activities in SN4741 cell, a dopaminergic neuronal cell, unlike wild type DJ-1. These findings suggest that DJ-1 regulates its structure and activities by concerted oxidative modifications of three cysteine residues. These studies broaden the understanding of regulatory mechanisms of DJ-1 that operate under oxidative conditions.

biochemistry↗

bioRxiv2020

The nucleus serves as the pacemaker for the cell cycle

Mitosis is a dramatic cellular process that affects all parts of the cell. In Xenopus embryos and extracts it is driven by the activation of a bistable trigger circuit, whose various components are localized in the nucleus, centrosome, and cytoplasm. In principle, whichever cellular location has the fastest intrinsic rhythm should act as a pacemaker for the process. Here we followed tubulin polymerization and depolymerization in Xenopus egg extracts supplemented with demembranated sperm, and thereby identified locations where mitosis first occurred. We found that mitosis was commonly first initiated at sperm-derived nuclei and their accompanying centrosomes, and then spread outward in circular trigger waves. The cell cycle was [~]20% more rapid at the nucleus/centrosome-associated trigger wave sources than in the regions of the extract that appeared not to be entrained by trigger waves. Nuclei produced from phage DNA, which did not possess centrosomes, also acted as trigger wave sources, but purified centrosomes in the absence of nuclei did not. We conclude that the nucleus accelerates mitotic entry and propose that it acts as a pacemaker for cell cycle. One Sentence SummaryStudies in cycling Xenopus egg extracts show that mitosis first occurs in the nucleus and then spreads outward through the cytoplasm in circular trigger waves.

cell biology↗