Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Proximity Proteomics Revealed Dicer1 and Transporters as Client Proteins of Aha1

ABSTRACT: Co-chaperone Aha1 activates HSP90 ATPase to promote the folding of client proteins. However, the client proteins of Aha1 are largely unknown. By employing ascorbate peroxidase (APEX) based proximity labeling, we identified 32 proximity proteins of HSP90 that are modulated by genetic depletion of Aha1. Among them, Dicer1 is one of the top-ranked proteins, which were further confirmed by streptavidin pull-down followed by Western blot analysis, demonstrating the reliability of the approach. Flag pull-down result showed interactions between endogenous HSP90 and Dicer1 and Aha1. The Dicer1 level is regulated synergistically by Aha1 and HSP90. Maturation-dependent interaction results showed a preferential binding of Aha1 and HSP90 to nascently translated Dicer1. Reconstitution of Aha1-depleted cells with WT Aha1 restored Dicer1 level, while the HSP90-binding-defective E67K mutant exhibited partial restoration. Moreover, knockdown of Aha1 and inhibition of HSP90 can diminish the levels of mature miRNA, let-7b and mir-30a. Overall, our study uncovers, for the first time, Dicer1 and transporter proteins as clients of Aha1 and HSP90.

INSTRUMENT(S): Q Exactive Plus

ORGANISM(S): Homo Sapiens (human) Pan Troglodytes (chimpanzee)

TISSUE(S): Permanent Cell Line Cell

SUBMITTER: Yen-Yu Yang

LAB HEAD: Yinsheng Wang

PROVIDER: PXD028980 | Pride | 2022-11-19

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	evidence.txt	Txt
	proteinGroups.txt	Txt
	shAha1-1.raw	Raw
	shAha1-2.raw	Raw
	shAha1-3.raw	Raw

Items per page:

1 - 5 of 11

Similar Datasets

c-Abl Dependent Tyrosine Phosphorylation of theCo-chaperone Aha1 Regulates Interaction with Hsp90

Project description:The ability of Heat Shock Protein 90 (Hsp90) to hydrolyze ATP is essential for its chaperone function. The co-chaperone Aha1 stimulates Hsp90 ATPase activity tailoring the chaperone function to specific “client” proteins. The intracellular signaling mechanisms directly regulating Aha1 association with Hsp90 remain unknown. Here we show that c-Abl kinase phosphorylates Y223 in human Aha1 (hAha1) promoting its interaction with Hsp90. This also increases Hsp90 ATPase activity,enhancesHsp90 interaction with kinase clients,and compromises the chaperoning of non-kinase clients such as glucocorticoid receptor and CFTR. Suggesting a new regulatory paradigm, we find that Y223 phosphorylation leads to ubiquitination and degradation of hAha1 in the proteasome. Finally pharmacologic inhibition of c-Abl prevents hAha1 interaction with Hsp90 thereby, hypersensitizing cancer cells to Hsp90 inhibitors bothin vitro and ex vivo.

2015-08-06 | PXD001737 | Pride

c-Abl Dependent Tyrosine Phosphorylation of theCo-chaperone Aha1 Regulates Interaction with Hsp90

2017-03-28 | MSV000080713 | MassIVE

A specialized Hsp90 co-chaperone network regulates steroid hormone receptor response to ligand

Project description:Heat shock protein-90 chaperone machinery is involved in the stability and activity of its client proteins. The chaperone function of Hsp90 is regulated by co-chaperones and post-translational modifications. Although structural evidence exists for Hsp90 interaction with clients, our understanding of the impact of Hsp90 chaperone function towards client activity in cells remains elusive. Here, we dissected the impact of newly identified co-chaperones in higher eukaryotes, FNIP1/2 (FNIPs) and Tsc1, towards Hsp90 client activity. Our data show that Tsc1 and FNIP2 form mutually exclusive complexes with FNIP1 and that unlike Tsc1, FNIP1/2 interact with the catalytic residue of Hsp90. Functionally, these co-chaperone complexes increase the affinity of the steroid hormone receptors glucocorticoid receptor and estrogen receptor to their ligands in vivo. Here, we provide a model for the responsiveness of the steroid hormone receptor activation upon ligand binding as a consequence of their association with specific Hsp90:co-chaperone subpopulations.

2023-05-30 | PXD030486 | Pride

Molecular determinants of Hsp90 recognition of Src kinase revealed by deep mutational scanning

Project description:Protein tyrosine kinases are involved in regulating growth and proliferation in cells and are often hyperactive in cancerous tissue. Tyrosine kinase inhibitors have been used to limit hyperactivity but become ineffective due to the appearance of resistance mutations. A common trait of hyperactive protein kinases is its strict client relationship with molecular chaperone Hsp90. However, the mechanism behind Hsp90 client kinase recognition is poorly understood. Here we measure the functional effect of thousands of single amino acid variants in the Src kinase domain to identify positions invovled in Hsp90 client recognition.

2023-05-22 | GSE218190 | GEO

Post-translational Regulation of FNIP1: a Rheostat for the Molecular Chaperone Hsp90

Project description:The molecular chaperone Hsp90 stabilizes and activates client proteins. Co-chaperones and post-translational modifications tightly regulate Hsp90 function and consequently lead to activation of clients. However, it is unclear whether this process occurs abruptly or gradually in the cellular context. We show that casein kinase-2 phosphorylation of the co-chaperone Folliculin-interacting protein-1 (FNIP1) on priming serine-938 and subsequent relay phosphorylation on serine-939, 941, 946, and 948 promotes its gradual interaction with Hsp90. This leads to incremental inhibition of Hsp90 ATPase activity and gradual activation of both kinase and non-kinase clients. We further demonstrate that serine/threonine protein phosphatase-5 (PP5) dephosphorylates FNIP1, allowing addition of O-GlcNAc (O-linked N-acetylglucosamine) to the priming serine-938. This process antagonizes phosphorylation of FNIP1, preventing its interaction with Hsp90, and consequently promotes FNIP1 lysine-1119 ubiquitination and proteasomal degradation. These findings provide a mechanism for gradual activation of the client proteins through intricate crosstalk of post-translational modifications of the co-chaperone FNIP1.

2019-01-31 | PXD012229 | Pride

Activation of autophagy depends on Atg1/ULK1 mediated phosphorylation of the chaperone Hsp90

Project description:The molecular chaperone Hsp90 is involved in the stability and activity of its client proteins which largely comprise of kinases. Phosphorylation of Hsp90 by these kinase clients provides a reciprocal regulatory mechanism between these proteins, however the mechanism of regulating the cellular pathway remains elusive. Here, we show that the serine/threonine kinase Atg1(yeast)/ULK1(mammalian) phosphorylates a conserved serine in the amino-domain of Hsp90 and reduces its ATPase activity hence lowers chaperone function. Broadly this modification negatively impacts the chaperoning of the kinase clients including Atg1/ULK1, yet enhances the activity of the non-kinase clients such as the heat shock factor and the steroid hormone receptors. Interestingly, ATG1/ULK1 mediated phosphorylation of the Hsp90 is essential for initiation of autophagy since yeast expressing a non-phosphorylatable Hsp90 were unable to undergo autophagy and the phosphomimetic Hsp90 mutants were underwent autophagy even in the absence of stimulus. Our findings provide a new paradigm where kinase client mediated phosphorylation of Hsp90 not only regulates the chaperone function but it is essential to initiate and regulate the relevant signaling pathway.

2023-09-08 | PXD038924 | Pride

The IMiD receptor CRBN determines quality control of transmembrane proteins

Project description:The complex architecture of transmembrane proteins requires quality control (QC) for folding and membrane positioning, a prerequisite for cellular homoeostasis and intercellular communication. However, it has remained unclear whether transmembrane proteins-specific QC hubs exist. Here we specify Cereblon (CRBN), the target of immunomodulatory drugs (IMiDs), as a co-chaperone that specifically determines chaperone activity of HSP90 towards transmembrane proteins by means of counteracting AHA1. This function is abrogated by IMiDs, which disrupt the interaction of CRBN with the closed conformation of HSP90. Cell surface proteomics identifies different transmembrane protein clients of the CRBN-AHA1-HSP90 nexus. Among these, we characterize the amino acid transporters LAT1 and CD98hc as determinants of IMiD activity in multiple myeloma (MM) both in vitro and in vivo. These data establish the CRBN-AHA1-HSP90 axis in the biogenesis of transmembrane proteins, link IMiD activity to tumor metabolism, and specify CD98 as an attractive diagnostic and therapeutic target in multiple myeloma.

2021-02-18 | PXD018244 | Pride

Effects of a phosphomimetic mutation and nucleotide binding on the interaction of Hsp90a with co-chaperone Aha1 analyzed by chemical cross-linking with mass spectrometry

Project description:Complex conformational dynamics are essential for the chaperone function of heat shock protein 90 (Hsp90), including transient, ATP-biased N-domain dimerization establishing ATPase competence. Biochemical data demonstrate that the intrinsic, but weak, ATP hydrolyzing activity of Hsp90 is markedly enhanced by the co-chaperone Aha1. However, cellular concentration of Aha1 is substoichiometric relative to Hsp90. In cells, interaction of this important co-chaperone with Hsp90 is up-regulated by posttranslational modifications (PTMs), including phosphorylation of a highly conserved tyrosine (Y313 in Hsp90a). Here we use chemical cross-linking with mass spectrometry to explore the the impacts of a phosphomimetic mutation (Y313E) and binding of a non-hydrolyzable ATP analog (AMP-PNP),on the structure Hsp90a and its interaction with Aha1.

2019-06-17 | PXD013476 | Pride

Mechanism of client kinase dephosphorylation by the HSP90-CDC37-PP5 complex

Project description:Activation of client protein kinases by the HSP90 molecular chaperone system is affected by phosphorylation at multiple sites on HSP90, on the kinase specific co-chaperone CDC37, and the client itself. Removal of regulatory phosphorylation from client kinases and release from the HSP90-CDC37 system depends on a Ser/Thr phosphatase PP5, which associates with HSP90 via its N-terminal TPR domain. Here we present the cryoEM structure of the oncogenic client kinase BRAFV600E bound to HSP90-CDC37, and structures of complexes of PP5 with that. Together with proteomic analysis of its phosphatase activity, our results reveal how PP5 is activated by recruitment to HSP90 complexes to dephosphorylate client proteins.

2023-03-11 | PXD033678 | Pride

Mechanism of client kinase dephosphorylation by the HSP90-CDC37-PP5 complex

2023-03-11 | PXD035934 | Pride