Cell Lysate Microarray for Mapping the Network of Genetic Regulators for Histone Marks

Abstract

SUMMARYProteins, as the major executer for cell progresses and functions, its abundance and the level of post-translational modifications, are tightly monitored by regulators.Genetic perturbation could help us to understand the relationships between genes and protein functions. Herein, to explore the impact of the genome-wide interruption on certain protein, we developed a cell lysate microarray on kilo-conditions (CLICK) with 4,837 knockout (YKO) and 322 temperature-sensitive (ts) mutant strains of yeast (Saccharomyces cerevisiae). Taking histone marks as examples, a general workflow was established for the global identification of upstream regulators. Through a single CLICK array test, we obtained a series of regulators for H3K4me3, which covers most of the known regulators in S.accharomyces. We also noted that several group of proteins that are involved in negatively regulation of H3K4me3. Further, we discovered that Cab4p and Cab5p, two key enzymes of CoA biosynthesis, play central roles in histone acylation. Because of its general applicability, CLICK array could be easily adopted to rapid and global identification of upstream protein/enzyme(s) that regulate/modify the level of a protein or the posttranslational modification of a non-histone protein.

Reverse-Phase Protein Microarrays for Overexpressed Escherichia coli Lysates Reveal a Novel Tyrosine Kinase

“…The RPPA can be used to identify a specific protein expression in all the lysates that are printed on the array . The RPPA has been used to profile the histone modifications in the nuclear extracts or cell lysates. , Since the cell lysates on the RPPA array can serve as the substrates, we propose that the overexpression of a kinase may increase the phosphorylation of the substrates and thus can be screened for kinases. In our study, we constructed RPPA with lysates of 4126 different overexpressed strains from the ASKA library and named E.…”