Functional redundancy is certainly a pivotal mechanism that supports the robustness

Functional redundancy is certainly a pivotal mechanism that supports the robustness of biological systems at a molecular cellular and organismal level. network and their functional significance in cultured cells and p38 signaling module with p38a and p38c XL184 playing more peripheral auxiliary functions. We also present the first evidence demonstrating that an evolutionarily conserved complex of p38b with glycogen synthase links stress XL184 sensing to metabolic adaptation. INTRODUCTION The p38 mitogen-activated protein kinase (MAPK) pathway is usually a key evolutionarily conserved XL184 mediator of an organism’s response to nerve-racking environmental stimuli (reviewed in recommendations 13 and 31). Similar to other members of the canonical MAPK family p38 kinases are activated as part of a conserved three-tier kinase cascade by dual phosphorylation of the threonine and tyrosine residues in the activation loop. Active p38 kinases phosphorylate a wide spectrum of both nuclear and cytoplasmic substrates including a limited number of subordinate kinases (e.g. MAPK-activated protein kinase 2 [MK2] p38-regulated/activated protein kinase [PRAK] and mitogen and stress-activated kinase 1 [MSK1] and MSK2) thus extending the cascade past the MAPK level (reviewed in reference 34). Through its substrates p38 regulates a range of cellular processes such as transcription (Mef2A [10] Mef2C and ATF1 -2 and -6) cell cycle (cyclin D1 and cyclin-dependent kinase [CDK] inhibitors) proliferation (epidermal growth factor receptor [EGFR] and fibroblast growth factor receptor 1 [FGFR1]) protein turnover (Siah2) and apoptosis (p53 and Bcl-2) (reviewed in recommendations 12 and 40). Given the level and intricacy from the causing regulatory network it isn’t surprising that the consequences of p38 signaling differ drastically based on cell type mobile microenvironment and the type and length of time of difficult stimuli. Heterogeneity from the p38 signaling network turns into even more complex because so many cells express several person in the p38 family members suggesting the chance of both redundant and exclusive upstream regulatory systems and downstream substrate private pools. In mammals the p38 family members includes 4 genes coding for p38α -β -δ and -γ. p38α mRNA can undergo alternate splicing that produces additional isoforms of the kinase including Mxi2 (5) Exip (49) and CSBP1 (26). Gene-targeting studies in mice are typically interpreted to suggest some degree of functional compensation XL184 between LIPG the four p38 kinases. Indeed mice transporting null alleles of p38β (1) -γ (36) and -δ (36 44 lack very easily discernible phenotypes suggesting that most if not all crucial functions of these kinases can be carried out by ubiquitously expressed p38α. Nonetheless evidence for the unique activities of various p38 isoforms is also beginning to build up. For instance p38γ appears to antagonize differentiation of satellite cells during adult skeletal muscle mass regeneration (20). Conversely p38α is usually a well-established activator of the myogenic differentiation program in a variety of cellular contexts including normal development (23) and differentiation of rhabdomyosarcoma cells (33). Clearly a systems biology approach would provide a more complete view of redundant and unique functions of p38 family kinases within a given cellular context. We sought to examine whether protein-protein conversation (PPI) mapping using affinity purification coupled to mass spectrometry (AP-MS) could yield initial data of sufficient resolution to define the extent of the regulatory network(s) in this family of closely related kinases. We chose to test this approach using as a well-defined model XL184 system. AP-MS has been successfully used to build proteome-scale conversation maps in yeast ((22). However large-scale data units often lack the resolution necessary to strongly establish conversation preferences within families of closely related proteins. The travel p38 module is much simpler compared to its mammalian counterpart. The family consists of three highly homologous proteins p38a -b and -c and only the first two users are believed to be bona fide p38 kinases. Coding sequences of all three kinases are contained within a single exon eliminating the possibility of the added complexity of alternatively spliced variants. Previous genetic analyses strongly established the functions of p38a and p38b in stress response and antimicrobial immunity (6 11 More recent studies began.

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