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Biology subjects

Yi, H.-W.

Publications and source records attributed to Yi, H.-W..

3 recordsLinked to original sources

Elevated Ubiquitin Phosphorylation by PINK1 Contributes to Proteasomal Impairment and Promotes Neurodegeneration

Ubiquitin (Ub), a central regulator of protein turnover, can be phosphorylated by PINK1 (PTEN-induced putative kinase 1) to generate S65-phosphorylated ubiquitin (pUb). Elevated pUb levels have been observed in aged human brains and in Parkinsons disease, but the mechanistic link between pUb elevation and neurodegeneration remains unclear. Here, we demonstrate that pUb elevation is a common feature under neurodegenerative conditions, including Alzheimers disease, aging, and ischemic injury. We show that impaired proteasomal activity leads to the accumulation of sPINK1, the cytosolic form of PINK1 that is normally proteasome-degraded rapidly. This accumulation increases ubiquitin phosphorylation, which then inhibits ubiquitin-dependent proteasomal activity by interfering with both ubiquitin chain elongation and proteasome-substrate interactions. Specific expression of sPINK1 in mouse hippocampal neurons induced progressive pUb accumulation, accompanied by protein aggregation, proteostasis disruption, neuronal injury, neuroinflammation, and cognitive decline. Conversely, pink1 knockout mitigated protein aggregation in both mouse brains and HEK293 cells. Furthermore, the detrimental effects of sPINK1 could be counteracted by co-expressing Ub/S65A phospho-null mutant but exacerbated by over-expressing Ub/S65E phospho-mimic mutant. Together, these findings reveal that pUb elevation, triggered by reduced proteasomal activity, inhibits proteasomal activity and forms a feedforward loop that drives progressive neurodegeneration.

biochemistry↗

Ubiquitin phosphorylation accelerates protein aggregation and promotes neurodegeneration in the aging brain

Ser65-phosphorylated ubiquitin (pUb) was found elevated in neurons of aged and neurodegenerative brains. Yet little is known whether a causative link exists between pUb level and brain aging. Here we show that the knockout of pink1, a Ub kinase, abolished pUb elevation and decelerated protein aggregation in aged mouse brains and cells with proteasomal inhibition. Conversely, over-expression of PINK1 but not the kinase-dead version increased the pUb level and accelerated protein aggregation by suppressing of proteasomal degradation. Furthermore, PINK1 over-expression in mouse hippocampus neurons increased pUb level and protein aggregation, slowly leading to mitochondrial injury, neurodegeneration, and cognitive impairment. Notably, the neuronal damages induced by PINK1 were rescued by the dominant negative Ub/S65A mutant, while Ub/S65E phosphomimetic mutant caused neuronal death. Together, an incidental increase of Ub phosphorylation can progressively and cumulatively cause the decline of Ub-dependent proteasomal activity, consequenting promotes neurodegeneration in the aging brain.

cell biology↗

Structural basis for the recognition of K48-linked Ub chain by proteasomal receptor Rpn13

The interaction between K48-linked ubiquitin (Ub) chain and Rpn13 is important for proteasomal degradation of ubiquitinated substrate proteins. Only the complex structure between the N-terminal domain of Rpn13 (Rpn13NTD) and Ub monomer has been characterized, and it remains unclear how Rpn13 specifically recognizes K48-linked Ub chain. Using single-molecule FRET, here we show that K48-linked diubiquitin (K48-diUb) fluctuates among three distinct conformational states, and a preexisting compact state is selectively enriched by Rpn13NTD. The same binding mode is observed for full-length Rpn13 and longer K48-linked Ub chain. Using solution NMR spectroscopy, we have solved the complex structure between Rpn13NTD and K48-diUb. In the structure, Rpn13NTD simultaneously interacts with proximal and distal Ub subunits of K48-diUb that remain associated in the complex, thus corroborating smFRET findings. The proximal Ub interacts with Rpn13NTD similarly as the Ub monomer in the known Rpn13NTD:Ub structure, while the distal Ub binds to a largely electrostatic surface of Rpn13NTD. Thus, a charge reversal mutation in Rpn13NTD can weaken the interaction between Rpn13 and K48-linked Ub chain, causing accumulation of ubiquitinated proteins. Moreover, blockage of the access of the distal Ub to Rpn13NTD with a proximity attached Ub monomer can also disrupt the interaction between Rpn13 and K48-diUb. Together, the bivalent interaction of K48-linked Ub chain with Rpn13 provides the structural basis for Rpn13 linkage selectivity, which opens a new window for modulating proteasomal function.

biophysics↗