Phosphorylation of (Ser 291) in the linker insert of Syk negatively regulates ITAM signaling in platelets
Creators: name: Dangelmaier, Carol
name: Vari, Hymavathi Reddy
name: Vajipayajula, Dhruv N
name: Elzoheiry, Manal
name: Wright, Monica
name: Iyer, Ashvin
name: Tsygankov, Alexander Y
name: Kunapuli, Satya P
Publisher: name: Taylor & Francis
Year: 2024
Subjects: subject: Biochemistry
subject: Microbiology
subject: FOS: Biological sciences
subject: Cell Biology
subject: Genetics
subject: Physiology
subject: Science Policy
subject: Immunology
subject: FOS: Clinical medicine
subject: Developmental Biology

description: Receptor-induced tyrosine phosphorylation of spleen tyrosine kinase (Syk) has been studied extensively in hematopoietic cells. Metabolic mapping and high-resolution mass spectrometry, however, indicate that one of the most frequently detected phosphorylation sites encompassed S297 (S291 in mice) located within the linker B region of Syk. It has been reported that Protein kinase C (PKC) phosphorylates Syk S297, thus influencing Syk activity. However, conflicting studies suggest that this phosphorylation enhances as well as reduces Syk activity. To clarify the function of this site, we generated Syk S291A knock-in mice. We used platelets as a model system as they possess Glycoprotein VI (GPVI), a receptor containing an immunoreceptor tyrosine-based activation motif (ITAM) which transduces signals through Syk. Our analysis of the homozygous mice indicated that the knock-in platelets express only one isoform of Syk, while the wild-type expresses two isoforms at 69 and 66 kDa. When the GPVI receptor was activated with collagen-related peptide (CRP), we observed an increase in functional responses and phosphorylations in Syk S291A platelets. This potentiation did not occur with AYPGKF or 2-MeSADP, although they also activate PKC isoforms. Although there was potentiation of platelet functional responses, there was no difference in tail bleeding times. However, the time to occlusion in the FeCl<sub>3</sub> injury model was enhanced. These data indicate that the effects of Syk S291 phosphorylation represent a significant outcome on platelet activation and signaling <i>in vitro</i> but also reveals its multifaceted nature demonstrated by the differential effects on physiological responses <i>in vivo</i>. <b>What is the context</b>Spleen tyrosine kinase (Syk) is present a number of cells and important in controlling the functions of various cells and organs.Syk is known to exist in two isoforms Syk L (long form or Syk A) and Syk S (short form or Syk B).It is known that phosphorylation events regulate Syk activation and activity.In several inflammatory disease conditions, Syk mutants are known to play a role.Phosphorylation of the Syk residue Serine 291 is known to occur, but its function in the regulation of Syk activation or activity is not known. Spleen tyrosine kinase (Syk) is present a number of cells and important in controlling the functions of various cells and organs. Syk is known to exist in two isoforms Syk L (long form or Syk A) and Syk S (short form or Syk B). It is known that phosphorylation events regulate Syk activation and activity. In several inflammatory disease conditions, Syk mutants are known to play a role. Phosphorylation of the Syk residue Serine 291 is known to occur, but its function in the regulation of Syk activation or activity is not known. <b>What is new</b>In this study, we generated a mutant mouse Syk S291A, which cannot be phosphorylated on serine residue. We evaluated the function of platelets isolated from these mice and compared them to platelets isolated from wild type littermates.We observed that the mutation in Syk L unexpectedly caused Syk S to disappear from a number of tissues.Platelet functions are enhanced in mutant mouse platelets compared to those from wild-type mice. In this study, we generated a mutant mouse Syk S291A, which cannot be phosphorylated on serine residue. We evaluated the function of platelets isolated from these mice and compared them to platelets isolated from wild type littermates. We observed that the mutation in Syk L unexpectedly caused Syk S to disappear from a number of tissues. Platelet functions are enhanced in mutant mouse platelets compared to those from wild-type mice. <b>What is the impact</b>These studies enhance our understanding of the impact of Serine 291 phosphorylation on the function of Syk in platelets. These studies enhance our understanding of the impact of Serine 291 phosphorylation on the function of Syk in platelets.