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© Research
Project

The interconnection between chromatin, transcription and splicing

Scientific Fields
Diseases
Organisms
Applications
Technique
Starting Date
11
Sep 2015
Status
Ongoing
Members
1
Structures
1

About

During the last several years, numerous publications have reported observations about the coupling between chromatin remodelling and splicing regulation. However, the mechanism of this interconnection remains elusive. Based on biochemistry and cellular biology, I have developed a new cell free system, which mimics coupling between these machineries. This system has demonstrated that chromatin influences the efficiency of intron removal in the course of gene expression through the regulation of U2snRNP splicing factor. Now, I aime at exploring deeper the interconnection between these machineries, using our in vitro transcription-splicing system and mass-spectrometry. The main experimental technique used to evaluate the role of histone PTMs on gene expression is chromatin immunoprecipitation (ChIP). This methodology can readily provide information on histone PTMs present in vivo, however it has a number of technical restrictions. For example, the detection of histone PTMs by ChIP can be done only if specific antibodies are available and the technique requires full accessibility of the antigen  to  the antibody. Moreover, the majority of available antibodies targets only one modification on a given amino acid, thus any combination of histone PTMs in an antigen will affect dramatically its recognition. To date in mammalian cells at least twelve types of PTMs have been reported at over 60 different amino acid residues within histones. Very recently, the number of identified histone modifications has increased by approximately 70%, and now totals 130. This progress was possible due to the application of advanced mass-spectrometry analysis of PTMs. Therefore, we have for project and challenge to decipher histone PTMs and characterize the composition of spliceosome when chromatin is transcribed using our cell free systems and mass spectrometry.