ABSTRACT
Nuclear hormone receptor coregulator-interacting factor 1 (NIF-1) is a zinc finger nuclear protein that was initially identified to enhance nuclear hormone receptor transcription via its interaction with nuclear hormone receptor coregulator (NRC). NIF-1 may regulate gene transcription either by modulating general transcriptional machinery or remodeling chromatin structure through interactions with specific protein partners. We previously reported that the cytoplasmic/nuclear localization of NIF-1 is regulated by the neuronal Cdk5 activator p35, suggesting potential neuronal functions for NIF-1. The present study reveals that NIF-1 plays critical roles in regulating neuronal morphogenesis at early stages. NIF-1 was prominently expressed in the nuclei of developing rat cortical neurons. Knockdown of NIF-1 expression attenuated both neurite outgrowth in cultured cortical neurons and retinoic acid (RA)-treated Neuro-2a neuroblastoma cells. Furthermore, activity-induced Ca(2+) influx, which is critical for neuronal morphogenesis, stimulated the nuclear localization of NIF-1 in cortical neurons. Suppression of NIF-1 expression reduced the up-regulation of neuronal activity-dependent gene transcription. These findings collectively suggest that NIF-1 directs neuronal morphogenesis during early developmental stages through modulating activity-dependent gene transcription.
Subject(s)
Intracellular Signaling Peptides and Proteins/metabolism , Neurites/physiology , Nuclear Proteins/metabolism , Animals , Calcium/metabolism , Cell Enlargement , Cell Line, Tumor , Cell Nucleus/drug effects , Cell Nucleus/metabolism , Cells, Cultured , Central Nervous System Agents/pharmacology , Cerebral Cortex/cytology , Cerebral Cortex/growth & development , Cerebral Cortex/physiology , DNA-Binding Proteins , Mice , Neurites/drug effects , Neurogenesis/drug effects , Neurogenesis/physiology , Rats , Transcription Factors , Transcription, Genetic/drug effects , Transcription, Genetic/physiology , Tretinoin/pharmacologyABSTRACT
Cardiotoxin analogue III (CTX III) and cobrotoxin (CBTX) isolated from the Taiwan cobra venom (Naja naja atra) are structurally homologous, small molecular weight, all-beta-sheet proteins, cross-linked by four disulfide bonds at identical positions. The conformational stabilities of these toxins are compared based on temperature-dependent chemical shifts and amide proton exchange kinetics using two-dimensional NMR spectroscopy. The structure of CTX III is found to be significantly more stable than that of CBTX. In both the toxins, beta-strand III appears to constitute the stability core. In CTX III, the stability of the triple-stranded beta-sheet domain is observed to be markedly higher than the double-stranded beta-sheet segment. In contrast, in CBTX, both structural domains (double- and triple-stranded beta-sheet domains) appear to contribute equally to the stability of the protein. Estimation of the free energy of exchange (Delta G(ex)) of residues in CBTX and CTX III reveals that the enhanced stability of the structure of CTX III stems from the strong interactions among the beta-strands constituting the triple-stranded beta-sheet domain and also the molecular forces bridging the residues at the N- and C-terminal ends of the molecule.
Subject(s)
Cobra Cardiotoxin Proteins/chemistry , Cobra Neurotoxin Proteins/chemistry , Animals , Circular Dichroism , Deuterium , Elapidae , Hot Temperature , Kinetics , Nuclear Magnetic Resonance, Biomolecular , Protein Conformation , Protein Folding , Protein Structure, Secondary , Solutions , ThermodynamicsABSTRACT
The effect(s) of TFE (2,2,2-trifluoroethanol) on three different conformational states (native, denatured, and carboxymethylated) of CTX III and RNase A has been examined. Contrary to the general belief, the results of the present study reveal that TFE can induce helical conformation in a protein which has no sequence propensity to form a helix. It is found that the helix induction in TFE is intricately related to the destabilization of the tertiary structural conformation in proteins. More importantly, the disulfide bonds in proteins are found to have significant influence on the TFE-mediated helix induction. The results obtained in this study strongly suggest that information pertaining to the influence of disulfide bonds on helix induction need to be considered to improve the accuracy of secondary structure prediction algorithms.
