Biology 6151

Critical Analysis of Kaneda Paper

Professor:  Dr. Uhde-Stone

Student:  Eric Wasiolek

Date:  November 7, 2005

Summary

Kaneda et al. develop a screening method (DCS - differential chromatin scanning)  to identify DNA fragments wrapped around histone proteins which are subject to acetylation/deacetylation.  It has long been known that acetylation/deacetylation of histones is one of the epigenetic mechanisms invovled in unravelling the DNA tightly packed in chromatin sufficiently for DNA polymerase and RNA polyermase II and associated proteins to access the DNA for replication and transcription.  Without these epigenetic mechanisms, including acetylation and deacetylation, it has been shown in certain cases, neither replication nor transcription can occur.  Kaneda et al identify many of the genes (DNA segments) targeted by acetylation.  Among their findings, some of the genes were cancerous, most of the genes were subject to acetylation/deacetylation mechanisms, and many exhibited differential expression (transcription activity) attributable to the level of histone acetylation.

Background On Revelant Biology:

DNA which may be as long as 280 million base pairs in a single human chromosome (or 10 centimeters in length), has to be tremendously condensed to fit into a chromosome.  This condensation is accomplished in large part by winding the DNA around a histone protein.  It is known that the total mass of histones in chromosomes is about equal to the total mass of DNA.  In addition, the histone/DNA complex, known as a nucelosome, is involved in many higher levels of folding and condensation to achieve the compaction required by the chromosome.  Because of this compaction, it is difficult or impossible for proteins such as DNA polymerase or RNA polymerase and associated proteins to access the DNA to perform replication or transcription, respectively.  Hence some unfolding of the chromosome structure is required at the point of origin and replication start sites for replication or transcription needs to take place.  This unfolding is performed by several epigenetic mechanisms, including acetylation of the histones.  The specific mechanism of the histone acetylation seems to be that the positive charge of lysines in the N-terminus (11-37 residues) of the histone are neutralized to eliminate their interaction with the negative charge of the phosphate group of the DNA.  The neutralized residues then extend the DNA/histone complex into a "beads on a string" form, which allows transcriptional and replicational access to the DNA.

Critical Analysis:

Although the paper focusses on the acetylation/deacetylation mechanism of providing access to DNA in chromatin, there are many other epigentic mechanisms which are involved in making DNA accessible to replication and transcription.  These include:  methylation of the lisines of the histones (this actually is a method analogous to deacetylation, it prevents acetylation allowing recondensation after transcription or replication has taken place), arginine side chains may be methylated, and there may be phosphorylation of serine and threonine residues of the histones.  Also, the addition of ubiquitin (a 76 amino acid protein) can have substantial influence on the chromosome structure.  None of these other mechanisms are discussed in any depth, nor is their interaction with acetylation/deacetylation discussed.  This would have made the paper a more meaningful exploration into the mechanisms behind chromosome unravelling and its effect on the transcription and replication processes.

It is good that the authors specifically identify which genes (DNA regions) seem to be acetylated, and give the accession numbers of these genes for further analysis, as well as the tester/driver DNA ratio. 

It was good that the authors identified cases where there was a profound activation in transcription level was induced by acetylation of a specific genetic region of a chromosome in a gastric cell cancer line.  It was further indicated that several genes involved in cell proliferation (common in cancers) were strongly affected by acetylation, indicating that regulation of acetylation might become a method for waging a battle against cell prolierations in cancers.  The authors specifically identified which genes these were that were involved in cell profileration.

It was also indicated by the authors that in some cases acetylation affects the transcription of multiple genes.  An obvious question to ask here, which the authors didn't seem to, was were these genes related, as in coregulated genes or genes involved in a common pathway or biological process?  Such information might have been secured by a simple bioinformatics search, or by comparison to known microarray data.

The authors also discovered that whereas HDAC targets were found throughout the genome, there were certain aceytlation/deacetylation hotspots (such as the 8q24.3 chromosomal region). 

Summary

In all I think the authors demonstrated the usefulness of their differential chromatin scanning technique to identifying acetylation/deacetylation regions of the genome, but more investigation of their findings should be performed, as should the whole area of developing drugs to affect acetylation of genomic regions involved in cell proliferation.  I still contend, that more discussion of other epigenetic mechanisms involved in the regulation of transcription on densely packed chromosomes should have been conducted, with a consideration of their interactions with a relation to acetylation mechanisms.