Molecular Biology Survival Guide for Chemists – IV Epigenetics

Cells in the body look incredibly different. Here are some pictures of a type of neuron found in the brain — the Purkinje cell http://www.google.com/images?client=safari&rls=en&q=purkinje+cell&ie=UTF-8&oe=UTF-8&oi=image_result_group&sa=X and several views of some liver cells — http://www.google.com/images?client=safari&rls=en&q=hepatocyte&ie=UTF-8&oe=UTF-8&oi=image_result_group&sa=X.

How can this be?   Back in the day, long before the human genome project, Dolly, and induced Pluripotent Stem Cells (iPSCs), I wondered if they didn’t simply jettison the parts of their genome they didn’t need.   This turned out to be true in a very limited sense for cells making antibodies, but the 3 billion or so basepairs of the Purkinje cell and the liver cell (hepatocyte) are identical.

We know that different cells express different parts of their genome.  Proteins are the best and longest studied genome products, but there is good evidence the microRNAs and other ‘noncoding’ (for protein) parts of the genome are also differentially expressed.   Large parts of the genome are essentially locked up by a set of proteins binding to specific regions of the genome.  This occurs on such a large scale that it was visible at the light microscopic level using dyes to stain it.   The locked up portion is called heterochromatin, the unlocked part is called euchromatin.  All very nice, but this just moves the problem one step back.  Why do different cells have different distributions of heterochromatin?

For any sort of explanation we must turn to epigenetics.  These are changes in DNA and/or the proteins which wrap it up so it fits in the cell which are not inherited from parent to child but which are inherited in some way from parent cell to child cell.  Just how this is done isn’t known completely, but there is no question that it occurs.

Presently we know of essentially two types of epigenetic change — modification of the DNA nucleotides themselves and modification of histones.

The easiest to understand is modification of cytosine, one of the 4 bases making up DNA.  A methyl group can be placed at the 5 position of cytosine — http://ndbserver.rutgers.edu/education/education_RNA.html.  Note that the 5 position is on the opposite side of cytosine from the side involved in base pairing.  This means that proteins binding to the double helix can get at it.  Since it adds bulk and mass to the outside of the DNA it also means that other proteins normally binding to cytosine can’t get at it.  In general, methylated cytosine in front of (5′ to) a protein coding gene, means it won’t be made into its gene product (a protein).

We have 3 enzymes which put methyl groups on cytosine at the 5′ position (DNMT1, 3a, 3b), and we even know how this is passed on from parent cell to daughter cell, when DNA is copied and copies given to the two daughter cells, but that would take us to far afield. The more introspective among you may wonder why all cells don’t have identical methylation patterns if this is so, and how differential cytosine methylation gets established.  The short answer is, we don’t really know.

There is another cytosine modification which is quite similar — hydroxymethyl cytosine.  We don’t know much, but it appears to be important for genome stability and a variety of other effects — X chromosome inactivation, imprinting anbd repression of repetitive genomic sequences (important as they constitute well over half of our genome).

The really big epigenetic effect are chemical modifications of histone proteins.  For a description of what they are see — https://luysii.wordpress.com/2010/04/16/the-cell-nucleus-and-its-dna-on-a-human-scale-iv/.  Actually there is a series of 5 posts on just how crowded and complicated things are in the nucleus.  For all 5 (probably soon to be 6) posts  see https://luysii.wordpress.com/category/the-cell-nucleus-on-a-human-scale/.

The skinny about them is that they are a way to compact DNA so it fits into the nucleus.  Our genome with its 3 billion basepairs actually has a length of 1 meter !   The thickness of the aromatic rings of the nucleotides is 3.4 Angstroms ( — 3.4 x 10^-10 meters). It must be mushed down to 10 microns (or 10^-5 meters).  8 histones form a squat cylinder called a nucleosome with a diameter of 110 Angstroms and height of 60 Angstroms.  147 nucleotides wrap around the nucleosome twice. The net effect is to shorten the overall length of DNA.  This results in DNA compaction by a factor of 10.

Histones are quite basic with lots of arginines and lysines which are positively charged at physiologic pH allowing them to counteract the negative phosphates holding DNA together.  They also have lots of serines and threonines in the parts of the nucleosome.

So what?  Histone proteins have been shown to have a large number of different chemical modifications.  Some are methylation. Lysine can have from 1 to 3 methyl groups, arginine 2.  One of the most important modifications is acetylation of the lysine amino group converting an amine to an amide with subsequent loss of basicity and its positive charge, so that it binds to the negatively charged phosphates less well.  This would loosen up DNA nucleosome binding, making DNA more accessible for transcription by the humungously sized RNA polymerase II complex.  For details, see The cell nucleus and its DNA on a human scale – VI — not written as of 15 Sep ’11 (but hopefully soon).

At least 10 different chemical modifications of histones are known.  The first 3 have been mentioned above.  The serines and threonines can be phosphorylated.  Some are ubiquitinated, some are sumoylated, some ADP ribosylated.  Then there is cis trans proline isomerization, di-iminization (e.g. arginine to citrulline) and NEDDylation.

The real complexity comes with just the first 3 (at least these are the best studied ones).  You will see the term’ histone tail‘ used to refer to the amino terminal or carboxy terminal ends protruding from the nucleosome cylinder, and not involved in binding DNA. This is where the modifications take place, and the combinatorial possibilities are enormous.  Of the amino terminal 36 amino acids of histone H3, half are modifiable — arg, lys, ser and thr — statistically it should be 20%.

Again, so what?  Phosphorylation is used all over the cell to determine which proteins bind and where they bind.  The proteins binding to the nucleosomes determine what can or can’t be done with DNA.  This has nothing to do with the nucleotide sequence of DNA, so it is epigenesis par excellance.

I leave it to your imagination (or your research project) how these changes are inherited when DNA is duplicated prior to mitosis. Somehow they are, but the mechanisms are unclear.  Somehow the 8 histones of the nucleosome must be disassembled, 8 more with similar modifications produced, to produce a nucleosome for each newly replicated DNA strand and its partner.

That’s plenty to digest for chemists unfamiliar with the material.  Persevere and you will be exposed to some incredibly elegant chemical and molecular biological gymnastics as the cell goes about its business.

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Comments

  • Panama corporation  On September 28, 2011 at 3:31 am

    ..When one looks at a genome sequence written out as a series of As Cs Gs and Ts or drawn as a map with the genes indicated by boxes on a string of DNA as in for example there is a tendency to imagine that all parts of the genome are readily accessible to the that are responsible for its expression. The DNA in the nucleus of a eukaryotic cell or the nucleoid of a prokaryote is attached to a variety of proteins that are not directly involved in genome expression and which must be displaced in order for the and other expression proteins to gain access to the genes. We know very little about these events in prokaryotes a reflection of our generally poor knowledge about the physical organization of the prokaryotic genome but we are beginning to understand how the packaging of DNA into chromatin influences genome expression in eukaryotes.

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