Autophagy︱Shen Hanming's group revealed a new mechanism of autophagy-related protein WIPI2 regulating mitochondrial outer membrane protein degradation and mitophagy

Written by ︱ Guang Lu, edited by Shen Hanming ︱ Sizhen Wang Autophagy is an evolutionarily highly conserved process in which cells degrade and recycle their own substances under external environmental changes or stimuli to maintain cell homeostasis and survival [1,2]
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Mitophagy is a selective autophagy that participates in the removal of damaged or dysfunctional mitochondria in cells and is a key mechanism of mitochondrial quality control [3], which has attracted much attention in recent years
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Abnormal or absent mitophagy is closely related to the occurrence and development of various human diseases, including tumors, neurodegenerative diseases or aging [4]
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 PINK1-Parkin-mediated mitophagy is the most well-studied mechanism so far
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Upon mitochondrial membrane depolarization, PINK1 rapidly accumulates in the mitochondrial outer membrane, and then recruits Parkin by phosphorylating ubiquitin on mitochondria and further phosphorylates to activate Parkin
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Activation of Parkin-mediated ubiquitination of mitochondrial outer membrane proteins forms a feed forward loop to further amplify the PINK1-Parkin-dependent mitophagy pathway [5]
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 WIPI2 is a key protein in autophagosome synthesis, which promotes the lipidation of LC3 and the formation of autophagosomes by recruiting the ATG12–ATG5-ATG16L1 complex [6,7]
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At present, the research of WIPI2 in non-selective autophagy is relatively in-depth, but its role in selective autophagy, especially mitophagy, is still poorly understood
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Professor Harper's research group from Harvard Medical School reported that WIPI2 is a key protein recruited to damaged mitochondria in the initial stage of mitophagy induction [8], and its specific function and regulatory mechanism still need to be further studied
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 On April 7, 2022, Professor Shen Hanming's team from the National University of Singapore/University of Macau published a research paper entitled "WIPI2 positively regulates mitophagy by promoting mitochondrial recruitment of VCP" in the journal Autophagy, explaining autophagy Positive regulation of related protein WIPI2 on mitophagy and its regulatory molecular mechanism
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Associate Professor Lu Guang of Sun Yat-Sen University School of Medicine and Dr.
Hayden Weng Siong Tan of the Department of Physiology, National University of Singapore are the co-first authors of this paper, and Professor Shen Hanming is the corresponding author of the paper
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Through CCCP-induced mitochondrial membrane depolarization, the authors found that WIPI2 was efficiently recruited to damaged mitochondria (Fig.
1B)
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Further experiments revealed that WIPI2 co-localized with key mitophagy cargo receptors including OPTN and NDP52 (Fig.
1E, G)
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These results (Figure 1) suggest that the recruitment of WIPI2 to mitochondria is an early event in mitophagy
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Figure 1 WIPI2 is recruited to damaged mitochondria during mitochondrial depolarization (Source: Lu G, et al.
, Autophagy, 2022) Using CRISPR-Cas9 technology, the authors constructed multiple WIPI2 knockout cells and found that WIPI2 knockout As a result, CCCP or OA-induced degradation of mitochondrial outer and inner membrane proteins was blocked (Fig.
2 AB)
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Using the mito-Keima approach, the authors further demonstrated that WIPI2 knockdown resulted in the inhibition of mitophagy (Fig.
2 CE)
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These results suggest that WIPI2 positively regulates mitophagy
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Figure 2 WIPI2 knockout inhibits mitophagy (Source: Lu G, et al.
, Autophagy, 2022) In order to further study the mechanism of WIPI2 regulating mitophagy, the authors found that VCP/p97 (valosin containing protein, containing valosin protein) and its cofactors such as UFD1 are key proteins that interact with WIPI2 during mitophagy induction
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Several studies have reported that VCP is recruited to damaged mitochondria during the induction of mitophagy, thereby mediating the degradation of ubiquitination-modified mitochondrial outer membrane proteins and the occurrence of mitophagy [9,10]
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The authors then verified the interaction of the two (Fig.
3 AC) and found that the N domain of VCP is required for its interaction with WIPI2 (Fig.
3 DE)
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The N domain of VCP is a key domain for its interaction with cofactors such as UFD1 and NPLOC4 [11], and the authors further verified the interaction between WIPI2 and the latter (Fig.
3F)
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These results suggest that WIPI2 interacts with the VCP-UFD1-NPLOC4 complex
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Figure 3 WIPI2 interacts with VCP-UFD1-NPLOC4 (Source: Lu G, et al.
, Autophagy, 2022) Since VCP is a key protein that mediates the degradation of mitochondrial outer membrane proteins, WIPI2 knockdown leads to mitochondrial outer membrane degradation Is the blocking related to VCP? The authors further found that VCP recruitment to mitochondria was impaired in WIPI2-knockout cells, and overexpression of WIPI2 reversed this phenotype (Fig.
4)
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These results suggest that WIPI2 is a key protein in the recruitment of VCP to mitochondria
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Figure 4 WIPI2 promotes the recruitment of VCP to mitochondria (Source: Lu G, et al.
, Autophagy, 2022) The last part of the article elaborates that WIPI2 is involved in cell fate selection
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During the initial stage of mitophagy, mitochondrial outer membrane protein degradation is accompanied by mitochondrial rupture and the release of pro-apoptotic factors [12]
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In WIPI2 knockout cells, mitochondrial outer membrane protein degradation was inhibited, thereby inhibiting the release of pro-apoptotic factors such as cytochrome C and the activation of other pro-apoptotic factors (Fig.
5 AB), thereby inhibiting mitochondrial autophagy.
phagocytosis-induced cell death (Figure 5 CE)
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 Figure 5 WIPI2 knockdown inhibits mitophagy-induced apoptosis (Source: Lu G, et al.
, Autophagy, 2022) Figure 6 Work summary: WIPI2 promotes mitophagy by recruiting VCP complexes (Source: Lu G, et al.
, Autophagy, 2022) article conclusion and discussion, inspiration and prospect In conclusion, this study demonstrated the positive regulation of WIPI2 on mitophagy through a variety of autophagy research methods and means
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At the molecular level, WIPI2 interacts with the VCP-UFD1-NPLOC4 complex and promotes its recruitment to damaged mitochondria, thereby mediating the ubiquitin-proteasome-dependent degradation of mitochondrial outer membrane proteins and the occurrence of mitophagy ( Figure 6)
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 In addition, this study suggests that WIPI2 is involved in the regulation of two major degradation systems (ie, ubiquitin-proteasome and autophagy-lysosome degradation pathways) in eukaryotic cells: on the one hand, WIPI2 regulates ubiquitin-proteasome-dependent degradation; On the one hand, WIPI2, as a key protein in autophagosome synthesis, is involved in autophagy-lysosome degradation
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This study expands the functional scope of WIPI2 and the understanding of the molecular regulatory mechanism of mitophagy
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 There are also some unanswered questions in this study
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For example, how is WIPI2 recruited to damaged mitochondria and what factors influence the interaction of WIPI2 with VCP and the recruitment of VCP to mitochondria? Does the recruitment of WIPI2 to VCP also exist in other cellular processes? What are the physiopathological implications of WIPI2-regulated mitophagy? The discussion of these series of questions will help to deepen the understanding of the function of WIPI2 and the molecular mechanism of mitochondria, and provide theoretical support for the prevention and treatment of mitophagy deficiency related diseases
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Link to the original text: https://doi.
org/10.
1080/15548627.
2022.
2052461 The first author, Associate Professor Lu Guang (left), the corresponding author Professor Shen Hanming (middle), and the co-first author Dr.
Hayden Weng Siong Tan (right) (photo courtesy of : Shen Hanming