PMA functions as an autophagy inhibitor through activation of a serine threonine kinase

Serine-threonine kinases have vital roles in various signalling pathways such as proliferation, differentiation and apoptosis. Thus, they served as crucial players in health and disease including cancer. Isozymes of these families show tissue specific expression patterns and function specifically in different pathways. Autophagy is another major process in the cell which degrades long-lived, non-functional organelles and proteins to sustain the homeostasis. Moreover, as well as serine threonine kinases, autophagy was also found to relate with cancer in different stages. To date, relationship between serine-threonine protein kinases and autophagy has not been well studied. Therefore, we utilized PMA which is one of the three different activatory signals of this kinase family to activate the certain subgroup and we studied the role of these kinases on autophagy regulation

Identification of a serine-threonine kinase as a novel autophagic regulator

Phospholipid dependent Serine/Threonine kinases are shown to be involved in cellular mechanisms and disease related pathways. Upon different intracellular stimuli, these kinases are activated and functions. Several chemical analogues such as Phorbol 12-Mystrate 13-Acetate (PMA) and Ceramide were synthesized to mimic intracellular stimuli to study function of these kinases. For several of these kinases, activation is dependent on both PMA and a Calcium ionophore such as ionomycin. Strikingly, deregulation of these kinases has been identified in several cancers. Recent studies showed that autophagy, which evolutionary conserved cellular degradation mechanism to maintain homeostasis, is also involved in carcinogenesis. According to literature, there are no robust studies to show the interaction between autophagy and serine threonine kinases. Thus, in our study we focused to identify a novel Ser/Thr kinases involved in regulation of autophagy in cancer

Involvement of autophagy in T cell biology

Autophagy is an essential cellular pathway that sequesters various cytoplasmic components, including accumulated proteins, damaged organelles or invading microorganisms and delivers them to lysosomes for degradation. The function of autophagy has been reported in various tissues and systems, including its role in the regulation of cellular immunity. Autophagy plays a fundamental role at various stages of T cell maturation. It regulates the thymocyte selection and the generation of T cell repertoire by presenting intracellular antigens to MHC class molecules. Autophagy is crucial for metabolic regulation of T cells, and therefore supports cell survival and homeostasis, particularly in activated mature T cells. Furthermore, deletion of specific autophagy-related genes induces several immunological alterations including differentiation of activated T cells into regulatory, memory or natural killer T cells. In this review, we emphasize the impact of autophagy on T cell development, activation and differentiation, which is pivotal for the adaptive immune system

Effect of serine/threonine kinases on autophagy mechanism

Autophagy is a degradation pathway, activating under stress conditions. It digests macromolecules, such as abnormal proteins and long-lived organelles by engulfing them and by subsequent delivery of the cargo to lysosomes. The members of the phospholipid-dependent serine/threonine kinases, involved in many signaling pathways, which are necessary for the regulation of cellular metabolic activation. Previous studies implicate that, serine/threonine kinases have crucial roles in the mechanism of many diseases depend on the activated and/or inactivated signaling pathway. Data indicates, the signaling pathways activated by serine/threonine kinases are also involved in activation of autophagy mechanism. However, the information about the effect of serine/threonine kinases on autophagy mechanism and the roles of these effects in disease formation is limited. In this study, we investigated the effect of activated serine/threonine kinases on autophagic pathway. We performed a commonly used autophagy technique, GFP-LC3 dot formation and by using microscopy analyses, we evaluated promotion and/or inhibition of autophagy in serine/threonine kinase-overexpressed fibroblasts as well as cancer cells. In addition, we carried out confocal microscopy analyses and examined autophagic flux by utilizing the differential pH sensitivities of RFP and GFP in mRFP-GFP-LC3 probe. Based on the shRNA-library based screening, we identified autophagy-related proteins affected by serine/threonine kinases. We further studied the involvement of serine/threonine kinases on the molecular mechanism of newly identified autophagy proteins and found that, autophagic pathway is indirectly controlled by serine/threonine kinases via specific autophagic proteins. Our data indicate the molecular connection between two critical cellular mechanisms, which have important roles in the formation of many disease pathologies, particularly cancer