Human DOK1 blocking peptide

Integrins are hetero-dimeric (alpha and beta subunits) type I transmembrane proteins that facilitate cell adhesion and migration. The cytoplasmic tails (CTs) of integrins interact with a plethora of intra-cellular proteins that are required for integrin bidirectional signaling. In particular, the beta CTs of integrins are known to recruit a variety of cytosolic proteins that often have overlapping recognition sites. However, the chronological sequence of beta CTs/cytosolic proteins interactions remains to be fully characterized. Previous studies have shown that the scaffold protein 14-3-3 zeta binds to phosphorylated beta CTs in activated integrins, whereas interactions of Dok-1 with phosphorylated beta CTs maintained integrins in the resting state. In this study, we examined the binding interactions between 14-3-3 zeta, Dok1, and phosphorylated integrin beta 2 and beta 3 CTs. We show that the scaffold protein 14-3-3 zeta interacts with the phosphotyrosine binding (PTB) domain of Dok1 even in the absence of the phosphorylated integrin beta CTs. The interactions were mapped onto the beta-sheet region of the PTB domain of Dok1. Furthermore, we provide evidence that the 14-3-3 zeta/Dok1 binary complex is able to bind to their cognate phosphorylated sequence motifs in the integrin beta CTs. We demonstrate that Thr phosphorylated pTTT beta 2 CT or pTST beta 3 CT can bind to 14-3-3 zeta that is in complex with the Dok1 PTB domain, whereas Ser phosphorylated beta 2 CT or Tyr phosphorylated beta 3 CT interacted with Dok1 in 14-3-3 zeta/Dok1 complex. Based on these data, we propose that 14-3-3 zeta/Dok1 complex could serve as a molecular switch providing novel molecular insights into the regulating integrin activation. (C) 2018 Elsevier Ltd. All rights reserved.

Dolichols are a class of isoprenoids that consist of highly polymerized and unsaturated long-chain isoprenes. They play crucial roles in protein glycosylation including N-glycosylation, because the oligosaccharide is assembled on a lipid carrier, dolichyl diphosphate. Arabidopsis DOLICHOL KINASE 1, AtDOK1 (At3g45040), encodes a functional dolichol kinase that is involved in plant reproductive processes. The expression of AtDOK1 is limited to highly pluripotent cells although protein glycosylation is thought to be required ubiquitously in the entire plant body. In this study, we further explored AtDOK1 functions by creating leaky knockdown mutants of DOK1. We used a microRNA-mediated gene suppression technique because knockout of DOK1 causes lethality. The DOK1 knockdown mutants showed an early flowering phenotype without any remarkable growth defect in vegetative tissues. Indeed, AtDOK1 was highly expressed in emerging shoot apical meristems as well as inflorescence and floral meristems. A subcellular localization study of DOK1 revealed that DOK1 was localized at the endoplasmic reticulum. Our findings suggest that the endoplasmic reticulum-localized catalytically active DOK1 is highly expressed in the meristems and is involved in the control of flowering time, possibly by post-transcriptional regulation including protein glycosylation.