Quality Control Mechanisms of Molecular Chaperones in the Folding and Degradation of Client Proteins

Molecular chaperones are essential proteins that assist in the folding of substrate ‘client’ proteins to adopt their functionally active three-dimensional structures. The process of protein folding in the cell occurs in a highly concentrated crowded cellular environment among various other macromolecules and amidst various cell stresses which result in issues of aberrant protein folding into toxic species and aggregates. Thus, to counteract these stressors, cells have evolved a complex network of chaperone proteins to maintain protein homeostasis, or proteostasis. Hsp70 is an essential molecular chaperone that acts on clients important for a wide variety of cellular functions. Hsp70 can facilitate refolding of clients to regain their function. However, it can also target client proteins to proteasomal degradation. Turnover of aberrantly folded or aggregation prone proteins such as tau implicates Hsp70 in various pathologies including neurodegenerative diseases. Another class of protein chaperones, termed ‘holdases’, act to delay protein aggregation. The small heat shock proteins (sHSP) systems possess such activity, binding to non-native conformations of clients. sHsps such as Hsp27 and αB crystallin exist as distributions of large oligomeric species that respond dynamically to pH and temperature stresses. Recent studies have demonstrated oligomeric rearrangements occur for sHsps to protect client proteins. A major outstanding question is how do these oligomeric assemblies’ complex structures sense cell stress or protein unfolding or aggregation. In addition to sensing cell stress, sHsps and holdase chaperones are also capable of bridging with the activities of other classes of chaperones, including the Hsp70 chaperone system. Hsp70 functions in concert with a network of co-chaperone proteins which diversify its protein folding capabilities. BAG3 is a nucleotide exchange factor (NEF) that facilitates the exchange of ADP and ATP in Hsp70. In addition, interactions with sHsp family chaperones have emerged, making it a promising target in elucidating the link between these two functionally distinct chaperone systems. The overall theme to my thesis work has been to characterize protein homeostasis achieved through pro-folding and pro-degradation pathways. A major focus of my thesis concerns the ability of Hsp70 to work in concert with the CHIP E3 ubiquitin ligase to target tau for polyubiquitination in a chaperone dependent manner, thus facilitating protein turnover. Another focus has been on a pro-folding function of chaperones, the so-called holdase function, where I have explored the structural rearrangements of the sHsp αB crystallin as well as another multifunctional chaperone, peroxiredoxin, and how these conformational changes and oligomeric rearrangements trigger with external stress and correlate with activation of chaperone activity. This thesis also explores the cooperation between sHsps and Hsp70 to xiii facilitate protein refolding, where I characterize rearrangements that occur in the Hsp27 oligomer distribution modulated by BAG3, and its implications on Hsp70 binding. One of the major techniques utilized in my thesis work is electron microscopy, obtaining structural information of protein complexes, from obtaining low resolution size distributions of sHsp oligomers to pushing resolution of Hsp70 in complex with CHIP beyond quaternary structural information to sub-nanometer resolution of the peroxiredoxin in its active chaperone form in complex with substrate. These studies serve as a foundation for future work on obtaining the structural basis of the decision process where chaperone proteins decide the fate of their client substrates.PHDBiological ChemistryUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttps://deepblue.lib.umich.edu/bitstream/2027.42/144085/1/orvvdom_1.pd

Photovoltaic Emulator LabVIEW Based Modeling and Simulation

The design goal for this project is to create a system capable of creating a consistent testing environment for photovoltaic inverters and charge controllers. The solution is to create a photovoltaic emulator which is a DC power supply that would have the same electrical output characteristics to a photovoltaic panel. In current systems, consistency problems arise when depending on nature to conduct tests on PV modules because it is impossible to properly replicate scenarios while testing specific conditions. This project will provide the user the ability to test their PV modules under various parameters which replicate real-world scenarios such as temperature and weather conditions. This paper will outline the design and simulation of a photovoltaic emulator. Regarding the change in learning environment due to Covid-19, the original scope of the project had to be appropriately adjusted. All aspects of “Section 2: Project Planning” as well as the “Appendix A: Senior Project Design Analysis” will still account for the original design choices made during fall quarter preparation. “Section 3: System Design” will more accurately reflect the true scope of project. Please refer to “Appendix B: Alterations Due to Covid-19” for more details on changes made to better accommodate the situation

Molecular and Electrophysiological Mechanisms Underlying Cardiac Arrhythmogenesis in Diabetes Mellitus.

This is the final version of the article. It first appeared from Hindawi via https://doi.org/10.1155/2016/2848759Diabetes is a common endocrine disorder with an ever increasing prevalence globally, placing significant burdens on our healthcare systems. It is associated with significant cardiovascular morbidities. One of the mechanisms by which it causes death is increasing the risk of cardiac arrhythmias. The aim of this article is to review the cardiac (ion channel abnormalities, electrophysiological and structural remodelling) and extracardiac factors (neural pathway remodelling) responsible for cardiac arrhythmogenesis in diabetes. It is concluded by an outline of molecular targets for future antiarrhythmic therapy for the diabetic population.GT was awarded a BBSRC Doctoral Training Award at the University of Cambridge for his PhD

Primary immune thrombocytopenia responding to antithyroid treatment in a patient with Graves’ disease

published_or_final_versionSpringer Open Choice, 21 Feb 201

Extranodal Natural-Killer/T-Cell Lymphoma, Nasal Type

The World Health Organization (WHO) classification recognizes 2 main categories of natural killer (NK) cell-derived neoplasms, namely, extranodal NK/T-cell lymphoma, nasal type, and aggressive NK-cell leukaemia. Extranodal nasal NK/T-cell lymphoma is more frequent in the Far East and Latin America. Histopathological and immunophenotypical hallmarks include angiocentricity, angiodestruction, expression of cytoplasmic CD3 epsilon (ε), CD56, and cytotoxic molecules and evidence of Epstein-Barr virus (EBV) infection. Early stage disease, in particular for localized lesion in the nasal region, is treated with chemotherapy and involved-field radiotherapy. On the other hand, multiagent chemotherapy is the mainstay of treatment for advanced or disseminated disease. L-asparaginase-containing regimens have shown promise in treating this condition. The role of autologous hematopoietic stem cell transplantation is yet to be clearly defined. Allogeneic hematopoietic stem cell transplantation, with the putative graft-versus-lymphoma effect, offers a potentially curative option in patients with advanced disease

Competing protein-protein interactions regulate binding of Hsp27 to its client protein tau.

Small heat shock proteins (sHSPs) are a class of oligomeric molecular chaperones that limit protein aggregation. However, it is often not clear where sHSPs bind on their client proteins or how these protein-protein interactions (PPIs) are regulated. Here, we map the PPIs between human Hsp27 and the microtubule-associated protein tau (MAPT/tau). We find that Hsp27 selectively recognizes two aggregation-prone regions of tau, using the conserved β4-β8 cleft of its alpha-crystallin domain. The β4-β8 region is also the site of Hsp27-Hsp27 interactions, suggesting that competitive PPIs may be an important regulatory paradigm. Indeed, we find that each of the individual PPIs are relatively weak and that competition for shared sites seems to control both client binding and Hsp27 oligomerization. These findings highlight the importance of multiple, competitive PPIs in the function of Hsp27 and suggest that the β4-β8 groove acts as a tunable sensor for clients

The Landscape of Long Non-Coding RNA Dysregulation and Clinical Relevance in Muscle Invasive Bladder Urothelial Carcinoma.

Bladder cancer is one of the most common cancers in the United States, but few advancements in treatment options have occurred in the past few decades. This study aims to identify the most clinically relevant long non-coding RNAs (lncRNAs) to serve as potential biomarkers and treatment targets for muscle invasive bladder cancer (MIBC). Using RNA-sequencing data from 406 patients in The Cancer Genome Atlas (TCGA) database, we identified differentially expressed lncRNAs in MIBC vs. normal tissues. We then associated lncRNA expression with patient survival, clinical variables, oncogenic signatures, cancer- and immune-associated pathways, and genomic alterations. We identified a panel of 20 key lncRNAs that were most implicated in MIBC prognosis after differential expression analysis and prognostic correlations. Almost all lncRNAs we identified are correlated significantly with oncogenic processes. In conclusion, we discovered previously undescribed lncRNAs strongly implicated in the MIBC disease course that may be leveraged for diagnostic and treatment purposes in the future. Functional analysis of these lncRNAs may also reveal distinct mechanisms of bladder cancer carcinogenesis

Structural basis for substrate gripping and translocation by the ClpB AAA+ disaggregase.

Bacterial ClpB and yeast Hsp104 are homologous Hsp100 protein disaggregases that serve critical functions in proteostasis by solubilizing protein aggregates. Two AAA+ nucleotide binding domains (NBDs) power polypeptide translocation through a central channel comprised of a hexameric spiral of protomers that contact substrate via conserved pore-loop interactions. Here we report cryo-EM structures of a hyperactive ClpB variant bound to the model substrate, casein in the presence of slowly hydrolysable ATPγS, which reveal the translocation mechanism. Distinct substrate-gripping interactions are identified for NBD1 and NBD2 pore loops. A trimer of N-terminal domains define a channel entrance that binds the polypeptide substrate adjacent to the topmost NBD1 contact. NBD conformations at the seam interface reveal how ATP hydrolysis-driven substrate disengagement and re-binding are precisely tuned to drive a directional, stepwise translocation cycle

The second IEEE international workshop on program debugging: IWPD 2011

published_or_final_versionProceedings of IEEE 35th Annual International Computer Software and Applications Conference Workshops (COMPSACW 2011), the 2nd IEEE International Workshop on Program Debugging (IWPD 2011), Munich, Germany, 18-22 July 2011, p. xlviii - xlvi