gene expression Flashcards
Gene expression
Gene expression is a consequence of RNA synthesis, processing, stability, and translation, as well as post-translational processing and protein stability. Examples of regulation can be found at each step.
Control of Transcription Initiation
transcription initiation is the MOST important mechanism for determining whether or not most genes are expressed, and consequently how much of which particular protein a specific cell contains.
DNA Control Elements
DNA elements that act locally. Binding of transcription factors to these elements controls expression of the gene that the element is associated with.
TATA box/Initiator sequence
a type of dna control element, generally 25-35 bps upstream of the transcription start site. It determines the site of transcription initiation and directs binding of RNA polymerase II. This is the site at which general transcription factors bind.
Promoter Proximal Elements
located within 200 bps upstream of transcription start site and are ~ 20 bps long, help to regulate transcription, and can be bound by factors in a cell type specific manner, a type of dna control element, effects the basal recruitment of transcriptional machinery
Enhancers
Contain multiple DNA control elements, each 8-20 bps in length, region of DNA that can be bound with proteins (activators) to activate transcription of a gene or genes. Thus, an entire enhancer can be 100-200 bps long. An enhancer can be 200-tens of kilobases upstream or downstream from the promoter or the last exon of the gene, or within an intron. Similar to promoter proximal elements, enhancers may help to regulate transcription in a cell type specific manner.
trans-acting element
is usually a DNA sequence that contains a gene. This gene codes for a protein (or microRNA or other diffusible molecule) that will be used in the regulation of another target gene. The trans-acting gene may be on the same chromosome as the target gene, but the activity is via the intermediary protein or RNA that it encodes. Trans-acting factors can switch from repressors to activators by recruiting either HATs or HDACs (ex-thyroid hormone receptor)
cis-Regulatory Elements (CREs)
are regions of non-coding DNA which regulate the transcription of nearby genes.
what are the two basic regulators of eukaryotic gene expression?
DNA control elements (cis) and transcriptional activators/ repressors (trans)
Thalassemias
An inherited anemia due to deficient production of b-globin protein by erythroid cells. Can occur due to different types of mutations- one of which can occur in the b-globin promoter (DNA control element), reducing the amount of b-globin mRNA and thus protein produced (is usually clinically mild).
Beta globin
a globin protein, which along with alpha globin (HBA), makes up the most common form of hemoglobin in adult humans, the HbA
Hemophilia B Leyden
Is an X-linked disorder that affects clotting. Affected males have 1% of normal factor IX active until puberty due to inherited mutations in a DNA control element in the promoter of the Factor IX gene (which prevents the binding of the appropriate transcriptional activators). Alternative transcriptional activators (which have an androgen receptor) can bind overlapping sites in the promoter, and at puberty it can bind at the promoter site and increase transcription such that males after puberty make ~60% the normal amount of factor IX.
Fragile X-Syndrome
Affects 1 in ~1500 males and results in mental retardation, dysmorphic facial features, and postpubertal macroorchidism. CGG repeat in the 5’ region of the FMR1 gene facilitates methylation of the cytosine residues in CpG islands and transcriptional inactivation of the FMR1 gene. Normal males have 6-50 or so CGG repeats in this region, however, affected males have an expansion of this repeat sequence (>200 copies)- leading to increased transcriptional silencing of the FMR1 gene.
Transcriptional Activators and Repressors
Proteins encoded by one gene that act on other genes to regulate their transcription. Can therefore diffuse around the nucleus and affect transcription of numerous genes. Can either activate or repress transcription. Multiple activators stimularte the transcription from both proximal and distal site. It is the combination of these protiens that lead to when, where, and how much of a protein is transcribed. the two classes inculde sepuence- specific DNA binding proteins and co-factors. Can recruit either histone acetyltransferases (HATS) or deacetylases (HDACs), eg if bound to hormone, changes conformation of TF.
Sequence-specific DNA binding proteins
bind to promoter or enhancer elements (DNA control elements) in their target genes to regulate transcription. The elements they bind to are usually 6-8 base pairs long. Usually bind DNA by inserting their a-helices into the major groove of DNA, making contacts between the amino acid side chains of the protein and the bases in the DNA . Contain two domains, DNA binding domain and activation/ repression domain, which allow you to mix and match (modularity)
DNA binding domain
The DNA binding domains (DBDs) are highly structured and evolutionarily conserved. They are folded so that they can “read” the DNA sequence and bind to their specific target DNA. Devides sequences- specific DNA binding proteins into different families, which have similar tertiary structure
homeodomain proteins
helix-turn-helix structure in which three alpha helices are connected by short loop regions. Many homeodomains induce cellular differentiation by initiating the cascades of coregulated genes required to produce individual tissues and organs. Members include Hox family, Pit1, Msx, etc.
Zinc-finger proteins
is characterized by the coordination of one or more zinc ions in order to stabilize multiple finger-like protrusions that make tandem contacts with their target molecule. Members include nuclear receptors such as estrogen receptor, androgen receptor, retinoic acid receptor
Basic leucine zipper proteins (bZIP)
contains a region that mediates sequence specific DNA binding properties and the leucine zipper that is required to hold together (dimerize) two DNA binding regions. The DNA binding region comprises a number of basic amino acids such as arginine and lysine. Members include c-fos and c-jun
Basic helix-loop-helix motif (bHLH)
is characterized by two α-helices connected by a loop. In general, transcription factors including this domain are dimeric, each with one helix containing basic amino acid residues that facilitate DNA binding. Members include MyoD, myogenin, Myf5
Craniosynostosis
Craniosynostosis is characterized by the premature closure of one or more sutures in the skull and affects 1/3000 infants. One particular variant of craniosynostosis (Boston-type) occurs as a result of a mutation in the homeodomain protein MSX2. MSX2 is normally required for proper craniofacial development by affecting the transcription of a number of genes important in this process. When the DNA binding domain (or homeodomain) of this protein has a one amino acid substitution, the protein binds DNA more strongly- giving a “gain of function” or “hypermorphic allele”. This mutated hyperactive protein then affects the transcription of other genes critical for suture closure, leading to craniosynostosis.
Androgen insensitivity syndrome (AIS)-
It occurs in males who are a normal karyotype (46 X,Y), but have mutations in either the DNA binding domain or the ligand binding domain of the androgen receptor (a zinc finger DNA binding protein). This makes the patients less responsive to androgens, leading to feminization or undermasculinization of the external genitalia at birth, abnormal secondary sexual development in puberty, and infertility. Depending on the degree to which the mutation disrupts the function of the androgen receptor- varying levels of AIS can be observed (complete, partial, mild).
Waardenburg Syndrome type II
Is characterized by deafness, pigmentation anomalies of the eyes, and other pigmentation defects (hair, skin). Mutations in the microphthalmia-associated transcription factor (MITF) gene (which encodes a bHLH DNA binding protein) are observed in 15-20% of the patients. This gene encodes a transcription factor that plays a major role in the development of melanocytes.
Dimerization of sequence specific DNA binding proteins
Many sequence specific DNA binding factors bind DNA as homo or heterodimers. The Zinc finger, bZIP, and bHLH can all form heterodimers. If each monomer of the heterodimer has a different DNA binding specificity, the formation of heterodimers will increase the number of potential sequences to which that family of sequence specific transcription factors can bind. “Combinatorial Control” can both regulate expression through new combinations of activation domains and change the DNA binding site
How do transcriptional activators or repressors, once bound to DNA control elements, stimulate transcription?
1) regulate assembly of initiation complexes and rate of initiation of transcription 2) regulate changes in chromatin structure influencing the ability of general transcription factors to bind to promoters
What are the two classes of chromatic- remodelling factos?
DNA-dependent ATPases (SWI/SNF)- disrupt histone octamers and DNA and F\factors that reversibly modify histones through acetylation (HATs and HDACs)