Malathi Narayan

Identification of Novel Cdc37 Interacting Proteins and Pathways in Human Alzheimers Disease Brain Tissue Using Mass Spectrometry

Alzheimer’s disease (AD) is the most common form of dementia and the 6th leading cause of death in the United States. The major pathological hallmarks observed in AD include the formation of intracellular neurofibrillary tangles comprised of phosphorylated forms of the microtubule associated protein tau, and the deposition of extracellular plaques composed of amyloid beta. Cdc37 is a co-chaperone of Hsp90, which recruits client kinases to the Hsp90 complex for folding and stabilization. It has been previously shown that Cdc37 can not only bind and preserve tau, but also stabilize kinases that can phosphorylate tau. The goal of the current study was to identify novel Cdc37- interacting proteins in human AD tissue compared to normal tissue using an immunoprecipitation-based approach combined with mass spectrometry. We identified 39 unique proteins that interacted with Cdc37 in AD samples only and 7 proteins that interacted with Cdc37 in normal samples only. 39 proteins were found to bind Cdc37 in both AD and normal tissue. Of these, 18 showed increased interaction in AD tissue, 10 showed increased interaction in normal tissue and 11 showed equal nteraction in both samples. Ingenuity Pathway Analysis of the data indicates that these Cdc37-interacting proteins could signal through the p70S6K, PI3K / Akt, TGFs, ErbB, NF- kB, calmodulin, p38 MAPK and JNK pathways. Identification of these novel proteins and pathways linked to Cdc37 may indicate its role both as a non-kinase co-chaperone and in other pathways in the AD brain.

Identification of Apo B48 and other novel biomarkers in amyotrophic lateral sclerosis patient fibroblasts

AIM Amyotrophic lateral sclerosis (ALS) is a debilitating fatal neurodegenerative disorder. 90-95% of ALS cases are sporadic with no clear risk factors associated with the disease. Identification of biomarkers associated with ALS may lead to early detection and make it more amenable to therapeutic intervention. MATERIALS & METHODS SILAC was used to quantitatively analyze the proteomes of ALS and control human fibroblasts. RESULTS Out of total of 861 proteins identified, 33 were found to be differentially regulated. ApoB48 and Hsp20 were downregulated, while Fibulin-1 was upregulated. CONCLUSION We report the differential regulation of these proteins in ALS fibroblasts, and their potential as novel biomarkers and possible drug targets for ALS.

Synthesis, Stereochemical Analysis, and Derivatization of Myricanol Provide New Probes That Promote Autophagic Tau Clearance

We previously discovered that one specific scalemic preparation of myricanol (1), a constituent of Myrica cerifera (bayberry/southern wax myrtle) root bark, could lower the levels of the microtubule-associated protein tau (MAPT). The significance is that tau accumulates in a number of neurodegenerative diseases, the most common being Alzheimers disease (AD). Herein, a new synthetic route to prepare myricanol using a suitable boronic acid pinacol ester intermediate is reported. An X-ray crystal structure of the isolated myricanol (1) was obtained and showed a co-crystal consisting of (+)-aR,11S-myricanol (2) and (-)-aS,11R-myricanol (3) coformers. Surprisingly, 3, obtained from chiral separation from 1, reduced tau levels in both cultured cells and ex vivo brain slices from a mouse model of tauopathy at reasonable mid-to-low micromolar potency, whereas 2 did not. SILAC proteomics and cell assays revealed that 3 promoted tau degradation through an autophagic mechanism, which was in contrast to that of other tau-lowering compounds previously identified by our group. During the course of structure-activity relationship (SAR) development, we prepared compound 13 by acid-catalyzed dehydration of 1. 13 had undergone an unexpected structural rearrangement through the isomyricanol substitution pattern (e.g., 16), as verified by X-ray structural analysis. Compound 13 displayed robust tau-lowering activity, and, importantly, its enantiomers reduced tau levels similarly. Therefore, the semisynthetic analogue 13 provides a foundation for further development as a tau-lowering agent without its SAR being based on chirality.

Data from SILAC-based quantitative analysis of lysates from mouse microglial cells treated with Withaferin A (WA).

Mass spectrometry data collected in a study analyzing the effect of withaferin A (WA) on a mouse microglial (N9) cell line is presented in this article. Data was collected from SILAC-based quantitative analysis of lysates from mouse microglial cells treated with either WA or DMSO vehicle control. This article reports all the proteins that were identified in this analysis. The data presented here is related to the published research article on the effect of WA on the differential regulation of proteins in mouse microglial cells [1]. Mass spectrometry data has also been deposited in the ProteomeXchange with the identifier PXD003032.

Cdc37: Implications in Regulation of Kinases and Proteins Linked to Neurodegenerative and Other Diseases

Abstract Autophagy is a cellular process that is important for removal and recycling of misfolded proteins and damaged cellular components, and maintenance of cellular homeostasis especially in neurons. Autophagic dysfunction has been reported in a number of other neurodegenerative diseases including Alzheimer’s disease (AD), Parkinson’s disease, lateral sclerosis (ALS), and Huntington’s disease. Cdc37, a co-chaperone that recruits a number of kinases to the Hsp90 complex for stabilization and folding, has been studied in the context of cancer. While Cdc37 has been widely studied as a co-chaperone of Hsp90, it is also a chaperone in its own right. Our work has shown that Cdc37 can stabilize tau and transactive response DNA-binding protein (TDP-43), which are proteins involved in AD, ALS, and other neurological disorders. Depletion of Cdc37 can lead to clearance of TDP-43 protein via autophagy. The Hsp90–Cdc37 complex has also been shown to regulate ULK1, a kinase that plays an integral role in autophagy. Additionally, a number of kinases including Akt, p38, and inositol-requiring enzyme 1, which are clients of Cdc37, have also been shown to regulate autophagy. Evidence for the role of Cdc37 in autophagy is starting to emerge, and understanding its function in autophagy in the context of neurodegenerative diseases may lead to its establishment as a drug target for clearance of toxic aggregates in AD and ALS.