Preliminary Research On The Interaction Between HELP3 And HELP4, YELP4, CTK1

Epigenetic modifications, particularly methylation and acetylation or other covalent histone modification, play an important role in regulating chromatin dynamics and therefore have a significant impact on gene expression. It has been revealed that yeast Elongator histone acetyltransferase complex facilitates RNAP II and takes part in the elongating process of GAL1 and SSA3 transcription.Human elongator complex has been isolated from HeLa cells and shown to be remarkably similar to its yeast counterpart in either components or functions. The most striking subunit of Elongator complex is the third subunit ELP3 which has histone acetyltransferase activity and belongs to GNAT superfamily. Deletion of ELP3 confers a set of phenotypes, some of which are associated with elongation defects. It is reported that ELP3 was also found to play a crucial role in neurobiology. The HAT activity of Elongator complex was found to acetylate histones, with specificity to H3 K14 and H4 K8.The C terminal HAT activity of ELP3 is essential for its function in vivo. Bioinformatics analysis showed that the N-terminal of ELP3 shares significant sequence homology to the Radical SAM superfamily.This domain contain the Fe4S4 cluster and can be binded to SAM.ELP4 is also a component of the Elongator complex, involved in transcription and tRNA modification. Elongator depletion results in the brain-specific downregulation of genes implicated in cell motility and migration. We hypothesize that a non-coding mutation in ELP4 impairs brain-specific Elongator-mediated interaction of genes implicated in brain development, resulting in susceptibility to seizures and neurodevelopmental disorders.Based on above results, this paper constrcted "Bait" fusion plasmid and"Prey"fushion plasmid pGADT7-Rect-yELP4,pGADT7-Rect-hELP4,pGADT7-Rect-CTK1,Divided them into three groups ,and then transformed them into yeast AH109 to yease two-hybind experiments verified the interaction between hElp3 protein with the other three proteins in vivo. Then verified the resuits of yease two-hybind experiments by GST pull-down experiments.The result of the experiment show that there is a interaction between hELP3 and hELP4, but the hELP3 subunit has no interaction with hELP4 and CTK1. The yeast two-hybrid experiment and pulldown experiment have confirmed this result.