Shenk uses an excerpt as his evidence that the molecules
that cling to the DNA are the ones that produce the "striking differences
between two organisms with the same genes" (178). Zimmer then goes on to
say that these "millions of proteins and other molecules" are the
things that determine which genes “produce transcripts and which cannot” (178).
Name some of types of molecules and proteins that inhibit or stimulate the
production of proteins to be transcripted during the process of protein
production. Also, include the process of the protein production as well as the
previously studied central dogma. How would these proteins possibly be affected
by the environment? Tie this back into the GxE theory of Shenk’s and what it would have to do with the differing traits in humans.
(Christina Li, christinali208@gmail.com)
Most scientists have long believed the sequence of nucleotides located on the alleles of our chromosomes is the deciding factor of our traits and phenotype. As it turns out, the there is actually another layer here, literally. There are "millions of proteins and other molecules" that cling to DNA and form the “epigenome – the packaging that surrounds the DNA” (Shenk 178, 158). These proteins and molecules play a large part in gene regulation in the transcription phase of protein production. Genes can be turned off by methylation. During methylation a –CH3 group is typically added to the nucleic acid cytosine. The reaction results in the DNA strand wrapping more tightly around the histone proteins which inhibits the availability of the gene to RNA polymerase and other transcription factors so gene expression is reduced. Histone acetylation is the process of adding a –COCH3 to the amino acids in the histone protein. This results in the genes attached to internal nuclear scaffolds that are typically not expressed to become loosened away from the histone protein and available to transcription factors. Now these genes are likely to be transcribed and expressed. For example, one experiment performed by Allfrey, Faulker, and Mirsky noted that “such modifications of histone structure, acetylation in particular, may affect the capacity of the histones to inhibit ribonucleic acid synthesis” (http://www.ncbi.nlm.nih.gov/pmc/articles/PMC300163/?page=1). This experiment proved the effect of acetylation promoted gene expression by adding acetate-2-C14 to calf thymus nuclei and that outside molecule beside the nucleotides can have an impact on gene expression. The DNA strands are studded with many other molecules, most of them still unknown and their complexity is still unfathomable to most. However, it is known that of the molecules “at least eight different classes have been characterized to date and many different sites have been identified for each class” (http://www.ncbi.nlm .nih.gov/pubmed/17320507). These molecules function in “disrupting chromatin contacts or by affecting the recruitment of nonhistone proteins to chromatin” (http://www.ncbi.nlm .nih.gov/pubmed/17320507). DNA is most often in the heterochromatin form so the strands are so highly condensed transcription is unlikely. However, chemical reactions with the studded molecules can alter the strands to become, at least briefly, euchromatin. Euchromatin is more loosely compact chromatin and genes are more likely to transcribed and expressed in this state since the genes are accessible to RNA polymerase and transcription factors. Although some specific identities of proteins are still questionable, it “been discovered that eukaryotic cells contain chromatin remodeling complexes, protein machines that use the energy of ATP hydrolysis to change the structure of nucleosomes temporarily so that DNA becomes less tightly bound to the histone core” (http://www.ncbi.nlm.nih.gov/books/NBK26834/). The whole key to gene expression is a structure and function application. These “are large protein complexes that can contain more than ten subunits” are hypothesized to be “used whenever a eukaryotic cell needs direct access to nucleosome DNA for gene expression, DNA replication, or DNA repair” (http://www.ncbi.nlm.nih.gov/books/NBK26834/). The protein complexes due to the molecular make-up change the shape of the DNA wound around the histone proteins, which makes them more or less available for transcription factors. This ultimately affects the expression of the affected genes. Another way to alter gene expression is through a process where “an enzymatically catalyzed covalent modification of the N-terminal tails of the four core histones”
ReplyDelete(continued)
ReplyDelete(http://www.ncbi.nlm.nih.gov/books/NBK26834/). Once again, the histone modification can loosen the DNA surrounding it which can increase gene expression since genes will be translated more often.
The central dogma states that the transcription of DNA leads to RNA which leads to proteins. David Shenk briefly describes the central dogma by stating “each gene initiates the process of assembling amino acids into proteins” (21). First RNA polymerase II attaches to the TATA box promoter located “upstream” from a gene. The binding of RNA polymerase is facilitated by transcription factors. RNA polymerase adds nucleotides in the 5’ to 3’ direction. It attaches an adenine to thymine, urine to adenine, guanine to cytosine, and cytosine to guanine. This transcript is now called pre-mRNA. Still in the nucleus, a guanine triphosphate cap is added to the 5’ end and a Poly-A tail is added to the 3’ end. The mRNA is also edited where non-coding regions called introns are excised and exons are spliced together by spliceosomes. Then the mRNA moves out of the nucleus into the cytoplasm. The guanine cap and Poly-A tail help the mRNA attach to the binding site of the ribosome. Protein synthesis is initiated by the start codon AUG on the mRNA. Anticodons that are complimentary to the codon read on the ribosome attach to the A-site of the ribosome. tRNAs have a corresponding amino acid attached to them by the enzyme aminoacyl-tRNA synthetase and ATP. During elongation of translation, codons, sets of three nucleotides, are read from the mRNA and complimentary anticodons bring the corresponding amino acids. The amino acids link together into a polypeptide by dehydration synthesis (forms peptide bonds) and kept in the P-site of the ribosome. The polypeptide chain terminates when the ribosome reads a stop codon. Then the protein continues to finalize its structure by secondary and tertiary structure. Then it is these proteins whether they are enzymes, transport mechanisms, etc. that make up who we are; “we are, each one of us, the sum of our proteins” (Shenk 22).
The environment as demonstrated in Shenk’s GXE model affects Gene expression. He explains “genes are constantly activated and deactivated by environmental stimuli, nutrition, hormones, nerve impulses, and other genes” (22). For example, currently the food we ate contains more hormones that it has in the past and this may be an explanation for girls going into puberty earlier. Not only can the environment affect gene expression, but also the protein itself. It was found in one experiment that a protein that formed alpha-helix secondary structure in one solvent actually formed beta-pleated sheets in a different solvent. The scientists therefore concluded that “environment is important in determining the secondary structure formed by an amino acid sequence; therefore schemes that predict secondary structure from amino acid sequence alone can never be totally successful” (http://www.pnas.org/content/89/10/4462.abstract).
A difference in structure between two identical amino acid sequences can mean completely different functions. The structure can affect the active site of an enzyme and the lock and key fit so either a reaction may not take place or different one will. This can be a difference that affects a trait. Proteins can also be affected by the environment due to changes in temperature and acidity. Enzymes will be denatured if the environment is too hot or too basic or acidic. Body environment in these ways can affect an enzymes ability to catalyze a reaction. Since these chemical reactions help to determine our traits, the environment can definitely affect our traits.
Lizzy Ettleson, lettleson@gmail.com
Because Lizzy focused more on histones and DNA organization, I am going to answer this question by using the example of a specific protein: the glucose transporter type 4 protein (GLUT4). When a person exercises, the muscles need glucose to perform cellular respiration to make ATP so that they have energy to contract. Insulin, a hormone released onto the bloodstream from beta cells in the pancreas, stimulates the uptake of glucose into skeletal muscle. By binding to the insulin receptor on the cell membrane of a muscle cell, insulin initiates a biological pathway within the cell that results in the glucose transporter type 4 protein (GLUT4) being translocated into the cell membrane as a transmembrane protein, through which glucose can enter the muscle cell by diffusion across a gradient (http://www.youtube.com/watch?v=KatbNCEBSDU). Because it has been shown that glucose cannot enter a muscle cell during exercise without the GLUT4 protein, it appears that “GLUT4 is essential for exercise-stimulated increases in muscle glucose uptake” (http://www.jbc.org/content/274/6/3253.short). Further evidence for the importance of GLUT4 in exercise is that transcription of the GLUT4 protein increases when muscles are active, and transcription “can be increased as much as two- to threefold after a few days of repeated exercise bouts” (http://www.ncbi.nlm.nih.gov/pubmed/15235326).
ReplyDeleteThe GLUT4 gene is transcribed into RNA in generally the same way as any other protein is transcribed in eukaryotes. First, the transcription factors must help RNA polymerase II bind to the promoter region of the gene. GLUT4 has two main transcription factors: myocyte enhancer factor 2A (MEF2A) and GLUT4 enhancer factor (GEF). It has been found that when only one of the transcription factors is present, RNA polymerase II will not bind or translate very much mRNA, but when both factors are present, they “increased promoter activity 4- to 5-fold” (http://www.pnas.org/content/100/25/14725.full). MEF2A and GEF are specific transcription factors, meaning that they lead to high levels of transcription only “at the appropriate time and place” (Campbell 359), which in this case are during exercise at the GLUT4 gene. The transcription factors recognize and bind to the TATA box in the promoter region, and then DNA polymerase II is able to bind to the promoter region to form the transcription initiation complex. The DNA strands then start to unwind and separate. RNA polymerase II moves in a 5' to 3' direction adding complementary nucleotides to the DNA template strand to form a complementary RNA strand. The RNA polymerase adds a cytosine base to complement a guanine base, guanine to complement cytosine, adenine to complement thymine, and uracil to complement adenine. As the RNA strand grows longer, it disconnects from the DNA template strand and the DNA rewinds. After the coding region ends, the DNA polymerase II reaches a polyadenylation signal that causes proteins to cut the pre-mRNA molecule off from the still transcribing polymerase so that it can be edited. A 5' cap is added to the 5' end of the mRNA that will later signal for a ribosome to attach there. A poly-A tail is added to the 3' end to stabilize the 3' end and to allow the mRNA to leave the nucleus. Then, the introns (sections that don't code for protein) are spliced out of the mRNA molecule by spliceosomes made up of snRNPs and other proteins (Campbell 333-337). The GLUT4 mRNA can be spliced in different ways. A few years ago, scientists “identified a novel splice variant of AS160 (variant 2 of AS160, AS160_v2) that lacks exon 11 and 12” (http://www.uniprot.org/citations/18771725). They found that the variant was more (cont next response)
Adele Padgett adele.padgett@gmail.com
active in people who had some insulin resistance than the full length protein. This is an example of alternative splicing, a process by which different exons can be reconnected after introns are spliced out of an mRNA molecule. Alternative splicing allows one gene, such as GLUT4, to produce many different proteins depending on how the exons are rejoined during splicing.
ReplyDeleteAfter editing, the GLUT4 mRNA leaves the nucleus and travels through the cytoplasm to ribosomes to be translated into a protein. First, a small ribosome subunit attaches to the 5' cap of the mRNA molecule. A initiator tRNA molecule with a UAC anticodon and an attached methionine amino acid binds to the AUG start codon on the mRNA. Then the large ribosomal subunit is attached to form the translation initiation complex with energy from GTP. When the initiation complex forms, the initiator tRNA is in the P site. Next, a tRNA with the anticodon complementary to the next codon in the mRNA enters the A site, carrying its corresponding amino acid. The rRNA helps form a peptide bond forms between the amino acids attached to the tRNAs in the P and A sites, forming the beginning of a chain. The chain disconnects from the tRNA in the P site, and then the tRNA in the P site shifts to the E site where it is released, and the tRNA in the A site shifts to the P site, making room for the next tRNA complementary to the next codon in the mRNA to enter the A site. Continuing this pattern and lengthening the polypeptide chain, the ribosome moves relative to the mRNA in a 5' to 3' direction until the stop codon, UAG, in the mRNA enters the A site of the ribosome. Instead of a tRNA with an amino acid, a release factor binds to the A site and causes the polypeptide to break off from the tRNA in the A site and exit the ribosome. As the GLUT4 polypeptide is produced, it begins to take on secondary structure (forming alpha helices and beta pleated sheets because of hydrogen bonding,) and tertiary structure (bending and folding because of hydrogen bonds, disulfide bridges, Van der Waals, and ionic interactions). Then it is either stored in vesicles until it is needed or translocated into the membrane of the cell if a person is exercising and insulin stimulates it to allow glucose to flood into a muscle cell.
When a healthy person exercises, GLUT4 gene expression increases producing more GLUT4 protein and allowing glucose to flood into muscle cells. However, when a person chooses not to exercise and to eat unhealthily, he or she is likely to develop diabetes type II, which involves developing an insulin resistance. This is a negative reaction with his or her environment. However, in response to the effects of the outer environment on the body's resistance to insulin, gene expression changes to produce the alternatively spliced variant of GLUT4 I mentioned earlier in my response. This variant is still able to translocate to the cell membrane during exercise even when muscle cell receptors are resistant to insulin. I, like Lizzy, relate this to the theme of structure and function because lacking the exons 11 and 12 causes GLUT4 to form a slightly different shape when it folds after translation, which in turn gives it a slightly different function than full length GLUT4.
Adele Padgett adele.padgett@gmail.com