Abstract
BackgroundTransgelin2, one of cytoskeletal actin binding proteins has recently been suggested to be involved in the formation of immune synapses. Although detailed function of transgelin2 is largely unknown, interactions between transgelin2 and actin appear to be important in regulating cellular functions of transgelin2. Because protein phosphorylation can change ability to interact with other proteins, comprehensive phosphorylation analysis of transgelin2 will be helpful in understanding its functional mechanisms.ResultsHere, a specific protein label-free quantitative phosphorylation analysis method combining immuno-precipitation, IMAC phosphopeptide enrichment technique and label-free relative quantification analysis was used to monitor the phosphorylation changes of transgelin2 overexpressed in Jurkat T cells under protein kinase C (PKC) and protein kinase A (PKA) activation conditions, two representative intracellular signalling pathways of immune cell activation and homeostasis. A total of six serine/threonine phosphorylation sites were identified including threonine-84, a novel phosphorylation site. Notably, distinct phosphorylation patterns of transgelin2 under the two kinase activation conditions were observed. Most phosphorylation sites showing specific kinase-dependent phosphorylation changes were discretely located in two previously characterized actin-binding regions: actin-binding site (ABS) and calponin repeat domain (CNR). PKC activation increased phosphorylation of threonine-180 and serine-185 in the CNR, and PKA activation increased phosphorylation of serine-163 in the ABS.ConclusionsMultiple actin-binding regions of transgelin2 participate to accomplish its full actin-binding capability, and the actin-binding affinity of each actin-binding region appears to be modulated by specific kinase-dependent phosphorylation changes. Accordingly, different actin-binding properties or cellular functions of transgelin2 may result from distinct intracellular signalling events under immune response activation or homeostasis conditions.Electronic supplementary materialThe online version of this article (doi:10.1186/s12953-015-0070-9) contains supplementary material, which is available to authorized users.
Highlights
Transgelin2, one of cytoskeletal actin binding proteins has recently been suggested to be involved in the formation of immune synapses
Our specific protein label-free phosphorylation quantitative analysis revealed distinct phosphorylation changes of transgelin2 in Jurkat T cell lines under the protein kinase C (PKC) and protein kinase A (PKA) activation conditions, which are comparable to immune cell activation and homeostasis conditions, respectively
Among the six transgelin2 phosphorylation sites identified in this study, four phosphorylation sites were located in three potential actin-binding regions: the actin-binding site (ABS), the calponin homology domain (CH) and the calponin repeat domain 1 (CNR1)
Summary
Transgelin, one of cytoskeletal actin binding proteins has recently been suggested to be involved in the formation of immune synapses. Research groups have reported somewhat contradictory results regarding the expression levels and cellular functions of transgelin2 Both oncogenic and tumor suppressive effects of transgelin have been observed depending on the types of cancer cell lines and tissue samples investigated. A quantitative phosphorylation analysis focusing on transgelin has not been previously conducted; several large-scale quantitative phosphoproteomics studies have reported phosphorylation level changes of transgelin in immune cells [17,18,19]. Because these studies investigated system-wide signal transduction pathways in different types of T cells, only one or two phosphorylation sites of transgelin were quantitatively analysed. To reveal the functional significance of transgelin phosphorylation events, comprehensive transgelin quantitative phosphorylation analysis under more specific kinase activation conditions is required
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.