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The BRISC deubiquitinating enzyme complex limits hematopoietic stem cell expansion by regulating JAK2 K63-ubiquitination.

ABSTRACT: Hematopoietic stem cell (HSC) homeostasis is controlled by cytokine receptor-mediated Janus kinase 2 (JAK2) signaling. We previously found that JAK2 is promptly ubiquitinated upon cytokine stimulation. Whether a competing JAK2 deubiquitination activity exists is unknown. LNK is an essential adaptor protein that constrains HSC expansion through dampening thrombopoietin (TPO)-induced JAK2 signaling. We show here that a LNK-associated lysine-63 (K63)-deubiquitinating enzyme complex, Brcc36 isopeptidase complex (BRISC), attenuates HSC expansion through control of JAK2 signaling. We pinpoint a direct interaction between the LNK SH2 domain and a phosphorylated tyrosine residue in KIAA0157 (Abraxas2), a unique and defining BRISC component. Kiaa0157 deficiency in mice led to an expansion of phenotypic and functional HSCs. Endogenous JAK2 and phospho-JAK2 were rapidly K63-ubiquitinated upon TPO stimulation, and this action was augmented in cells depleted of the BRISC core components KIAA0157, MERIT40, or BRCC36. This increase in JAK2 ubiquitination after BRISC knockdown was associated with increased TPO-mediated JAK2 activation and protein levels, and increased MPL receptor presence at the cell surface. In addition, BRISC depletion promoted membrane proximal association between the MPL receptor and pJAK2/JAK2, thus enhancing activated JAK2/MPL at the cell membrane. These findings define a novel pathway by which K63-ubiquitination promotes JAK2 stability and activation in a proteasome-independent manner. Moreover, mutations in BRCC36 are found in clonal hematopoiesis in humans. This research may shed light on the mechanistic understanding of a potential role of BRCC36 in human HSCs.

SUBMITTER: Donaghy R

PROVIDER: S-EPMC6450430 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

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The BRISC deubiquitinating enzyme complex limits hematopoietic stem cell expansion by regulating JAK2 K63-ubiquitination.

Donaghy Ryan R Han Xu X Rozenova Krasimira K Lv Kaosheng K Jiang Qinqin Q Doepner Miriam M Greenberg Roger A RA Tong Wei W

Blood 20190212 14

Hematopoietic stem cell (HSC) homeostasis is controlled by cytokine receptor-mediated Janus kinase 2 (JAK2) signaling. We previously found that JAK2 is promptly ubiquitinated upon cytokine stimulation. Whether a competing JAK2 deubiquitination activity exists is unknown. LNK is an essential adaptor protein that constrains HSC expansion through dampening thrombopoietin (TPO)-induced JAK2 signaling. We show here that a LNK-associated lysine-63 (K63)-deubiquitinating enzyme complex, Brcc36 isopepti ...[more]

PMID: 30755420

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Project description:Alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPARs) are the primary mediators for inter-neuronal communication and play a crucial role in higher brain functions including learning and memory. Our previous work demonstrated that AMPARs are subject to ubiquitination by the E3 ligase Nedd4, resulting in EPS15-mediated receptor internalization and Ubiquitin (Ub)-proteasome pathway (UPP)-dependent degradation. Protein ubiquitination is a highly dynamic and reversible process, achieved via the balance between ubiquitination and deubiquitination. However, deubiquitination of mammalian AMPARs and the responsible deubiquitinating enzymes remain elusive. In this study, we identify USP46 as the deubiquitinating enzyme for AMPARs. We find that AMPARs are subject to K63 type ubiquitination, and USP46 is able to deubiquitinate AMPARs in vivo and in vitro. In heterologous cells and neurons, expression of USP46 results in a significant reduction in AMPAR ubiquitination, accompanied by a reduced rate in AMPAR degradation and an increase in surface AMPAR accumulation. By contrast, knockdown of USP46 by RNAi leads to elevated AMPAR ubiquitination and a reduction in surface AMPARs at synapses in neurons. Consistently, miniature excitatory postsynaptic currents recordings show reduced synaptic strength in neurons expressing USP46-selective RNAi. These results demonstrate USP46-mediated regulation of AMPAR ubiquitination and turnover, which may play an important role in synaptic plasticity and brain function. Protein ubiquitination is a highly dynamic and reversible process, achieved via the balance between ubiquitination and deubiquitination. The glutamatergic AMPARs, which mediate most of the excitatory synaptic transmission in the brain, are known to be subjected to Nedd4-mediated ubiquitination; however, the deubiquitination process and the responsible deubiquitinating enzymes (DUBs) for mammalian AMPARs remain elusive. We find that AMPARs are subject to K63-type ubiquitination, and identify USP46 as the DUB for AMPARs. USP46 deubiquitinates AMPARs in vitro and in vivo. Up- or down-regulation of USP46 leads to changes in AMPAR ubiquitination, surface expression, and trafficking, as well as the strength of synaptic transmission. USP46-mediated regulation of AMPAR ubiquitination and turnover may play an important role in synaptic plasticity and brain function.

Dataset Information

The BRISC deubiquitinating enzyme complex limits hematopoietic stem cell expansion by regulating JAK2 K63-ubiquitination.

Publications

The BRISC deubiquitinating enzyme complex limits hematopoietic stem cell expansion by regulating JAK2 K63-ubiquitination.

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