Phosphorylation-Elicited Quaternary Changes of GA Binding Protein in Transcriptional Activation

Abstract

Enrichment of nicotinic acetylcholine receptors (nAChR) on the tip of the subjunctional folds of the postsynaptic membrane is a central event in the development of the vertebrate neuromuscular junction. This is attained, in part, through a selective transcription in the subsynaptic nuclei, and it has recently been shown that the GA binding protein (GABP) plays an important role in this compartmentalized expression. The neural factor heregulin (HRG) activates nAChR transcription in cultured cells by stimulating a signaling cascade of protein kinases. Hence, it is speculated that GABP becomes activated by phosphorylation, but the mechanism has remained elusive. To fully understand the consequences of GABP phosphorylation, we examined the effect of heregulin-elicited GABP phosphorylation on cellular localization, DNA binding, transcription, and mobility. We demonstrate that HRG-elicited phosphorylation dramatically changes the transcriptional activity and mobility of GABP. While phosphorylation of GABP␤ seems to be dispensable for these changes, phosphorylation of GABP␣ is crucial. Using fluorescence resonance energy transfer, we furthermore showed that phosphorylation of threonine 280 in GABP␣ triggers reorganizations of the quaternary structure of GABP. Taken together, these results support a model in which phosphorylation-elicited structural changes of GABP enable engagement in certain interactions leading to transcriptional activation.The neuromuscular junction (NMJ) is a specialized structure involved in communication between the motor neurons and skeletal muscle cells. As the NMJ forms, signals from the motor nerve terminal initiate a complex sequence of processes in the myonuclei, which result in the accumulation of a specific set of gene products in the postsynaptic membrane, such as the nicotinic acetylcholine receptors (nAChR), acetylcholine esterase, and utrophin (13, 44). However, in the absence of innervation, the muscle seems to be somewhat prepatterned (31,60).The present models for postsynaptic differentiation suggest that at least three different mechanisms operate in the polynucleated muscle cell. On the one hand, nerve-evoked electrical activity represses transcription of synaptic genes in extrasynaptic nuclei but on the other hand, several mechanisms may possibly contribute to the increased local concentration of nAChR at the NMJ. Two nerve-derived signaling molecules enhance both transcriptional and posttranslational mechanisms that govern clustering and targeting of the synaptic proteins directly underneath the motor nerve: (i) agrin released from the motor nerve terminal into the basal lamina of the synaptic cleft stimulates redistribution of previously unlocalized cell surface nAChR to the postsynaptic membrane domain (for reviews, see references 22 and 42), and (ii) another neurotrophic factor selectively augments transcription of nAChR genes in the subsynaptic nuclei (for reviews, see references 44 and 47). At present, the most likely candidate for the latter activity is the so-called acet...

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Nuclear respiratory factor 2 senses changing cellular energy demands and its silencing down-regulates cytochrome oxidase and other target gene mRNAs

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Basic helix–loop–helix factors recruit nuclear factor I to enhance expression of the NaV 1.4 Na+ channel gene

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