Abstract:Aggresome formation is a major strategy to enable cells to cope with proteasomal stress. Misfolded proteins are assembled into micro-aggregates and transported to the microtubule organizing center (MTOC) to form perinuclear aggresomes before their degradation through autophagy. So far, multiple factors have been identified as the activators of micro-aggregate formation, but much less is known about the regulatory mechanisms of their transport. Here, we report that proteasomal stress leads to the activation of … Show more
“…7), further assessing that MKK3-signaling pathway sustains pro-survival effects through the ERCC1-positive regulation. Overall results prompt to support that the MKK3/p38delta MAPK-signaling pathway likely sustains pro-survival effects through ERCC1-positive regulation in CRC cells and suggest that, in line with recent report 25 , the p38delta MAPK isoform represent a key molecular player also in the control of the autophagic process.Fig. 5MKK3 affects 5-FU efficacy via activation of p38delta MAPK. a , b Western Blot on protein lysates from sh/scr and sh/MKK3 HT29 and SW620 sublines ( a ) and sth/scr and sth/p38δ transfected HT29 and SW620 cells ( b ) either untreated or exposed to 5-FU.…”
Colorectal cancer (CRC) is one of the most common malignant tumors worldwide and understanding its underlying molecular mechanisms is crucial for the development of therapeutic strategies. The mitogen-activated protein kinase-kinase 3 (MKK3) is a specific activator of p38 MAP kinases (p38 MAPKs), which contributes to the regulation of several cellular functions, such as proliferation, differentiation, apoptosis as well as response to drugs. At present, the exact MKK3/p38 MAPK pathway contribution in cancer is heavily debated because of its pleiotropic function. In this work, we retrospectively explored the prognostic and pathobiologic relevance of MKK3 in a cohort of CRC patients and assessed MKK3 molecular functions in a panel of CRC lines and colonocytes primary cultures. We found increased MKK3 levels in late-stage CRC patients which correlated with shorter overall survival. Herein, we report that the MKK3 targeting by inducible RNA interference univocally exerts antitumor effects in CRC lines but not in primary colonocytes. While MKK3 depletion per se affects growth and survival by induction of sustained autophagy and death in some CRC lines, it potentiates response to chemotherapeutic drug 5-fluorouracil (5-FU) in all of the tested CRC lines in vitro. Here, we demonstrate for the first time that in CRC the MKK3 specifically activates p38delta MAPK isoform to sustain prosurvival signaling and that such effect is exacerbated upon 5-FU challenge. Indeed, p38delta MAPK silencing recapitulates MKK3 depletion effects in CRC cells in vitro and in vivo. Overall, our data identified a molecular mechanism through which MKK3 supports proliferation and survival signaling in CRC, further supporting MKK3 as a novel and extremely attractive therapeutic target for the development of promising strategies for the management of CRC patients.
“…7), further assessing that MKK3-signaling pathway sustains pro-survival effects through the ERCC1-positive regulation. Overall results prompt to support that the MKK3/p38delta MAPK-signaling pathway likely sustains pro-survival effects through ERCC1-positive regulation in CRC cells and suggest that, in line with recent report 25 , the p38delta MAPK isoform represent a key molecular player also in the control of the autophagic process.Fig. 5MKK3 affects 5-FU efficacy via activation of p38delta MAPK. a , b Western Blot on protein lysates from sh/scr and sh/MKK3 HT29 and SW620 sublines ( a ) and sth/scr and sth/p38δ transfected HT29 and SW620 cells ( b ) either untreated or exposed to 5-FU.…”
Colorectal cancer (CRC) is one of the most common malignant tumors worldwide and understanding its underlying molecular mechanisms is crucial for the development of therapeutic strategies. The mitogen-activated protein kinase-kinase 3 (MKK3) is a specific activator of p38 MAP kinases (p38 MAPKs), which contributes to the regulation of several cellular functions, such as proliferation, differentiation, apoptosis as well as response to drugs. At present, the exact MKK3/p38 MAPK pathway contribution in cancer is heavily debated because of its pleiotropic function. In this work, we retrospectively explored the prognostic and pathobiologic relevance of MKK3 in a cohort of CRC patients and assessed MKK3 molecular functions in a panel of CRC lines and colonocytes primary cultures. We found increased MKK3 levels in late-stage CRC patients which correlated with shorter overall survival. Herein, we report that the MKK3 targeting by inducible RNA interference univocally exerts antitumor effects in CRC lines but not in primary colonocytes. While MKK3 depletion per se affects growth and survival by induction of sustained autophagy and death in some CRC lines, it potentiates response to chemotherapeutic drug 5-fluorouracil (5-FU) in all of the tested CRC lines in vitro. Here, we demonstrate for the first time that in CRC the MKK3 specifically activates p38delta MAPK isoform to sustain prosurvival signaling and that such effect is exacerbated upon 5-FU challenge. Indeed, p38delta MAPK silencing recapitulates MKK3 depletion effects in CRC cells in vitro and in vivo. Overall, our data identified a molecular mechanism through which MKK3 supports proliferation and survival signaling in CRC, further supporting MKK3 as a novel and extremely attractive therapeutic target for the development of promising strategies for the management of CRC patients.
“…During proteasomal stress, UbK48-linked polyubiquitinated proteins accumulate and package to form aggresome, which are recognized by antibodies. UbK48-positive structures are co-localized with other aggresome markers, such as p62, HDAC6 and PINK1S (13,24,25). To examine the role of just synthesized proteins in aggresome formation, we studied aggresome formation efficiency in MG132-treated cells after blocking transcription by ActD or translation by ANI.…”
Section: Resultsmentioning
confidence: 99%
“…However, some misfolded proteins form dispersed aggregates rather than aggresomes during proteasome inhibition, such as HeLa, Huh7 and Cos7 (55,58). In contrast to aggresomes formed around MTOC, dispersed aggregates are distributed in the cells and do not result in cytoskeleton rearrangements, suggesting that dispersed aggregates may represent intermediate particles during aggresome assembly (24,58). The defects of aggresome formation in these cells may be caused by the lack of critical regulators of aggresome assembly, such as the subtype of p38 kinase (24).…”
Section: Discussionmentioning
confidence: 99%
“…In contrast to aggresomes formed around MTOC, dispersed aggregates are distributed in the cells and do not result in cytoskeleton rearrangements, suggesting that dispersed aggregates may represent intermediate particles during aggresome assembly (24,58). The defects of aggresome formation in these cells may be caused by the lack of critical regulators of aggresome assembly, such as the subtype of p38 kinase (24). Due to their biochemical and morphological similarities to the protein inclusion bodies in neurodegenerative diseases, aggresomes have received much attention in the past twenty years (41,59).…”
Aggrephagy, the aggresome-related protein degradation system, represents a protective cellular response to shuttle misfolded proteins into the microtubule-organizing center for degradation through the autophagic pathway during stress conditions, including heat shock, oxidative stress and proteasome inhibition. In response to proteasome failure, many genes are transcriptionally activated to facilitate ubiquitinated proteins to be cleared via the aggrephagy pathway. Although many regulators involved in aggresome formation have been identified, the mechanism how transcriptional activation promotes aggresome formation remains unknown. Here, we have demonstrated that nuclear factor erythroid 2-related factor 2 (Nrf2) accumulated in the nucleus and activated the transcription of sequestosome-1 (p62) during proteasome inhibition in 293 cells. Loss of Nrf2 resulted in failure of aggresome formation and cell death; whereas overexpression of p62 alleviated Nrf2 knockdown-induced aggresome formation defects and promoted cell survival. Notably, blocking Nrf2 activation using a p38/MAPK inhibitor prevented proteasome inhibitor-induced aggresome formation. These findings suggested that Nrf2 may be a critical regulator of aggresome formation, which protects cells from proteasome dysfunction-induced stress.
“…However, TAK1 may also affect p62 body formation through additional modifications since we found that another kinase (TBK1), known to phosphorylate p62 at both Ser‐349 and Ser‐403 , is dispensable for p62 body formation in response to pp242 treatment. It remains unclear whether TAK1 is the kinase directly responsible for p62 phosphorylation or whether it is carried out by kinases acting downstream of TAK1 (such as p38 MAP kinases ). Further study will also be necessary to determine how p62 scaffolds TAK1 activation.…”
The protein p62/Sequestosome 1 (p62) has been described as a selective autophagy receptor and independently as a platform for pro‐inflammatory and other intracellular signaling. How these seemingly disparate functional roles of p62 are coordinated has not been resolved. Here, we show that TAK1, a kinase involved in immune signaling, negatively regulates p62 action in autophagy. TAK1 reduces p62 localization to autophagosomes, dampening the autophagic degradation of both p62 and p62‐directed autophagy substrates. TAK1 also relocalizes p62 into dynamic cytoplasmic bodies, a phenomenon that accompanies the stabilization of TAK1 complex components. On the other hand, p62 facilitates the assembly and activation of TAK1 complexes, suggesting a connection between p62's signaling functions and p62 body formation. Thus, TAK1 governs p62 action, switching it from an autophagy receptor to a signaling platform. This ability of TAK1 to disable p62 as an autophagy receptor may allow certain autophagic substrates to accumulate when needed for cellular functions.
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