RACK1: a novel substrate for the Src protein-tyrosine kinase

Abstract

RACK1 is one of a group of PKC-interacting proteins collectively called RACKs (Receptors for Activated CKinases). Previously, we showed that RACK1 also interacts with the Src tyrosine kinase, and is an inhibitor of Src activity and cell growth. PKC activation induces the intracellular movement and co-localization of RACK1 and Src, and the tyrosine phosphorylation of RACK1. To determine whether RACK1 is a Src substrate, we assessed phosphorylation of RACK1 by various tyrosine kinases in vitro, and by kinase-active and inactive mutants of Src in vivo. We found that RACK1 is a Src substrate. Moreover, Src activity is necessary for both the tyrosine phosphorylation of RACK1 and the binding of RACK1 to Src's SH2 domain that occur following PKC activation. To identify the tyrosine(s) on RACK1 that is phosphorylated by Src, we generated and tested a series of RACK1 mutants. We found that Src phosphorylates RACK1 on Tyr 228 and/ or Tyr 246, highly-conserved tyrosines located in the sixth WD repeat that interact with Src's SH2 domain. We think that RACK1 is an important Src substrate that signals downstream of growth factor receptor tyrosine kinases and is involved in the regulation of Src function and cell growth.

RACK1 Regulates Directional Cell Migration by Acting on Gβγ at the Interface with Its Effectors PLCβ and PI3Kγ

“…We have shown previously that RACK1 can bind G␤␥ and selectively regulate its functions (Chen et al, 2004a), suggesting that RACK1 may impinge on cell migration by interaction with G␤␥. However, RACK1 is a multifunctional protein that has been shown to interact with many other proteins that are known to be involved in cell migration, such as integrin, PKC, Src, and ERKs (Schechtman and Mochly-Rosen, 2001;Chang et al, 2002;Vomastek et al, 2007). We therefore first evaluated if these RACK1-interacting proteins play a role in Jurkat cell migration.…”

RACK1 inhibits the serum‐ and anchorage‐independent growth of v‐Src transformed cells

“…We identified RACK1 as a Src substrate and an inhibitor of c-Src kinase activity and NIH 3T3 cell growth [7][8][9]. However, it is not yet known what influence RACK1 has on v-Src kinase activity or cell transformation.…”

Phosphorylation of RACK1 in plants

“…The study by Urano An important finding of the study by Urano et al (2015) 20 is that phosphorylation of RACK1 affects its protein stability. While the post-translational modification events have previously reported for mammalian RACK1 protein, 21,22 its impact on protein stability has not. The finding by Urano et al (2015) 20 promotes a new regulatory system in which the action of RACK1 is controlled by phosphorylation and subsequent protein degradation (Fig.…”

“…20 Phosphorylation of RACK1 has been previously reported in mammals. [21][22] However, a fundamental difference is that mammalian RACK1 is phosphorylated at tyrosine (Tyr) sites whereas plant RACK1 is phosphorylated at serine (Ser) and threonine (Thr) sites. Therefore, mammalian RACK1 is a substrate of tyrosine kinase whereas plant RACK1 is a substrate of serine/threonine kinase.…”

“…23 It should be noted that the Tyr phosphorylation site of mammalian RACK1 is also required for its binding to its Tyr kinase. Specifically, Tyr228 and Tyr246 are required for RACK1 binding to Src kinase 21 and Tyr52 of RACK1 is required for its binding to c-Abl kinase. 22 Urano et al (2015) 20 showed that Ser122 and Thr162 (Thr162 of RACK1A, Thr161 of RACK1B and RACK1C) of Arabidopsis RACK1 are phosphorylated by an atypical Ser/Thr kinase, WITH NO LYSINE 8 (WNK8).…”