Differential phosphorylation-based regulation of αB-crystallin chaperone activity for multipass transmembrane proteins

Abstract

We have previously shown that αB-crystallin (CRYAB), a small heat shock protein (sHsp) that prevents irreversible aggregation of unfolded protein by an ATP-independent chaperone activity, plays a pivotal role in the biogenesis of multipass transmembrane proteins (TMPs) assisting their folding from the cytosolic side of the endoplasmic reticulum (ER) (D'Agostino et al., 2013). Here we present evidence, based on phosphomimetic substitutions, that the three phosphorytable serine residues at position 19, 45 and 59 of CRYAB play a different regulatory role in this novel chaperone activity: S19 and S45 have a strong inhibitory effect, either alone or in combination, while S59 has not and counteracts the inhibition caused by single phosphomimetic substitutions at S19 and S45. Interestingly, all phosphomimetic substitutions determine the formation of smaller oligomeric complexes containing CRYAB, indicating that the inhibitory effect seen for S19 and S45 cannot be ascribed to the reduction of oligomerization frequently associated to a decreased chaperone activity. These results indicate that phosphorylation finely regulates the chaperone activity of CRYAB with multipass TMPs and suggest a pivotal role for S59 in this process.

Protective Effect of Rosamultin against H2O2‐Induced Oxidative Stress and Apoptosis in H9c2 Cardiomyocytes

“…Ser19 and ser45 have strong inhibitory effects alone or in combination, while ser59 does not have an inhibitory effect, and could offset the inhibitory effect of monophosphorimide in ser19 and ser45. The phosphorylation of ser19 and ser45 alone or in combination resulted in a significant decrease in cryAB chaperone activity, whereas the monophosphorimide in ser59 had no such effect [ 43 ]. After being pretreated with rosamultin, the level of cryAB decreased, and the level of pCryAB S59 expression increased.…”

αB-crystallin as a promising target in pathological conditions – A review

“…αB-crystallin has proved to play a crucial role in the biogenesis of multipass transmembrane proteins (TMPs) by assisting their folding from the cytosolic face of the endoplasmic reticulum (ER) [19]. Furthermore, Ciano et al (2016) established that phosphorylation of serine, resident at position 59, finely regulates the chaperone activity of HspB5 with studied TMPs [20]. Recent studies demonstrated the competence of HspB5 to interact with membrane channels.…”

Phosphorylation of the Chaperone-Like HspB5 Rescues Trafficking and Function of F508del-CFTR

“…However, a second hypothesis emerges from what occurs with two TMPs (Frizzled 4 receptor and ATP7B) similar to the multi-spanned transmembrane CFTR where HspB5 rescues the mutated TMPs. Thus, HspB5 could rescue F508del-CFTR rather than promoting its degradation [18,19].…”

“…HspB5 counteracts aggregation and rescues proper localization of mutated Frizzled 4 receptor and ATP7B, two mutated TMPs associated, respectively, with familial exudative vitreoretinopathy and Wilson's disease [18]. This beneficial effect on the misfolded TMPs was demonstrated to be regulated by HspB5 phosphorylation status [19]. In view of these data, we decided to evaluate if HspB5 could facilitate misfolded CFTR degradation, much like HspB1 and HspB4, or in contrast, promotes proper localization of mutated CFTR, as reported with other TMPs.…”

“…These data corroborate our choice of this cellular model to decipher the impact of HspB5 expression in CF. The ability of HspB5 to correct localization of TMPs at the PM was suggested to be dependent on phosphorylation [19]. HspB5 phosphorylation status was assessed using phosphoserine-specific antibodies that recognize the three known phosphoserines, Ser-19, Ser-45, and Ser-59 in HspB5.…”