Unit 8

Define

- Regulatory proteins- Regulate Transcription, Have two types

- Transcription factors- One type of regulatory protein, They bind directly to DNA sequence.

- Coregulators- Second type of regulatory protein, DO NOT bind directly to DNA sequence

- General Transcription Factors- One of two groups of transcription factors, Bind to the core promoter region of a gene (normally has the TATA box), Main function is to recruit RNA pol 2

- Specific (regulatory) Transcription Factors- Second group of transcription factors, Bind to specific DNA sequences, needed for cell specific and time specific gene expression, include activators and repressors which controls the rate at which transcription occurs

- Ehancers- Regulatory elements that increase rate of transcription, distinct from core promoter

- Proximal enhancers- Located close to the gene they regulate, they provide binding site for specific activator transcription factors, proteins bound directly to them can directlt interact with GTF and RNA pol II

- Distal enhancers- Located far from the gene they regulate, crucial for cell-type-specific and developmental gene expression, forms chromatin loop to bring enhancer and its bound activator proteins into physical contact

- Coactivators- Type of coregulator, aid in activating in transcription

- Corepressors- Type of coregulator, repress transcription

- Chromatin- Compacted DNA 10,000 fold in order to fit in certain places

- Canonical histones- H1, H2A, H2B, H3, and H4, diverge into core histones and linker histones.

- Core histones- Consist of the histones, H2A, H2B, H3 and H4, the function is to form a complex around DNA to form nucleosomes

- Linker histones- Its just H1, it serves as a link between histones

- Variant histones- incorporated into nucleosomes in DNA-rep AFTER replication, variants identifies in all histones except H4, the functions consist of epigenetic regulation, DNA replication, transcription, and chromosome segregation

- Nucleosome- Basic structural units of chromatin

- Epigenetic inheritance- Info stored in sequence of DNA is replicated and transferred to daughter cells

- Acetylation- Type of histone modification, when acetyl groups are added to histone tails, neutralizes the positive charge of histone tail

- Methylation- Type of histone modification, Methyl groups are added to specific amino acid residues on histone proteins, effects highly context-dependent and site-specific.

- Phosphorylation- Type of histone modification, transfer of phosphate (from ATP) to histone tail (adds negative charge)

- Ubiquitination- Last type of histone modification, attached ubiquitin to histones (generally 1 in histone modification)

- Nucleosome sliding- Type of chromatin remodeling, moves histone along DNA sequence.

- Nucleosome eviction/displacement- Type of chromatin remodeling, REMOVES a nucleosome from the DNA (the thing the DNA is wrapped around)

- Histone variant exchange- Last type of chromatin remodeling, swaps a core histone within a nucleosome with specialized histone variant

- Cellular memory- Type of epigenetic control of transcription, ability to maintain a specific gene expression pattern through cell divisions.

- Position-effect variegation- Type of epigenetic control of transcription, the expression of a gene is silenced when relocated to place adjacent to region of heterochromatin, also results in variegated phenotype

- Genome imprinting- Process in which certain genes are expressed in a parent-of-origin-specific manner (copies inherited from mom or dad expressed)

- X-chromosome inactivation- Occurs in female mammals to equalize gene dosage of x-linked gene between males and females (WOMEN ARE SILENCED)

Concepts

- Detail the GAL system as an example of Transcription factors (detail: What occurs when galactose is present, what occurs when galactose is not present, Gal4 protein, Gal3 and Gal80 proteins, domains of the Gal4 proteins)- Yeast make use of extracellular galactose by converting it into glucose. Abundance of galactose determines the level of transcription in the GAL metabolic pathway, Gal4 important, Gal4 is regulated by Gal80 and Gal3, Gal80 prohibits Gal4, Gal3 promotes Gal4. Gal80 is always on Gal4 but Gal3 comes in with the galactose and takes away Gal80 to start up Gal4.

- Detail the 3 structural domains of histone proteins- Histone fold- Mediates the interaction between histones to form heterodimers

Histone-fold extensions- responsible for protein-protein interactions

Flexible tails- Primary site of pots-translational modification

- Detail the nucleosome structure. What are the 2 primary function of nucleosomes- Composed of DNA (146 bp wrapped around histone octamers (2 copies of each core histone protein. The two functions are DNA compaction (reduces length of DNA by 6 or 7 fold) and Gene regulation (act as a physical barrier by restricting access to the DNA).

- Detail the “stages” of chromatin folding- 

Nucleosome- Compacts the DNA 6 or 7 fold, eukaryotic cells need 10,000 fold compaction

30-nm fiber- Nucleosome beads fold again into a more compact helical structure, H1 binds to linker DNA and pulls them closer together, compacts DNA approx 40-fold

Higher order folding- Chromatin undergoes even more folding to create large scale structure inside the nucleus

        Chromatin loops- 30nm fibers are organized into loops and tethered to protein         scaffold

        Chromosome territories- During interphase, each chromosome occupies a         specific, distinct region of the nucleus, prevents tangling

        Mitotic chromosomes- during mitosis, chromatin reaches highest level of         compaction, loops and domains supercoil and condense dramatically to form         the compact chromosomes

- Explain histone modification.- Its reversible chemical changes made to the histone proteins, has a code to signal wether gene should be active or inactive, primarily occurs on N-terminals tails of core histomes.

- Detail the following types of histone modifications

        - Acetylation- Acetyl groups are added to the histone tails which neutralizes the positive charge of the histone tail, weakens the DNA interactions and loosens the structure which increased accessablility

        - Methylation- Methyl groups are added to SPECIFIC amino acids residues on histone proteins, the effects are dependent on certain things, gene activation H3; recruits proteins that facilitates transcription, gene repression, H4; recruits proteins that increase compaction of chromatin.

        - Phosphorylation- Transfer of phosphate to histone tails, needs ATP, adds a negative charge to histone, which repels DNA (negative charge)

        - Ubiquitination- Attaching ubiquitin to histones; Activations H2B; acts as a mark that increases transcriptions of specific gene. Repressions H2A; marks the gene as “silent” NO Transcription.

- Detail chromatin remodeling- Chromatin remodeling is a process that alters the structure of chromatin which regulates gene expression, but requires ATP hydrolysis.

- Explain the 3 main mechanisms of chromatin remodeling

        - Nucleosome sliding- Sliding is when the remodeling complex moves the histone along the DNA sequence which exposes or hides specific regulatory DNA regions, and makes the DNA accessible to transcription factors and RNA pol II

        - Nucleosome eviction- Remodeling complex removes a nucleosome from the DNA which allows for the stable binding of transcription factors, Histone octamer can then be re-deposited elsewhere

        - Histone variant exchange- Chromatin remodelers can swap a core histone protein within a nucleosome with specialized histone variant which alters the ability of nucleosome, and once again increases accessibility of the DNA

- Detail the following types of epigenetic inheritance

        - Cellular memory- Remembering information and passing that on through subsequent cell divisions, stem cells are developed through differentiation which is remembered and passed down. DNA methylation and Histone Modification.

        - Position effect variegation (PEV)- Expression is silenced when it is relocated to a new position adjacent to a region of heterochromatin (tightly packed DNA which is transcriptionally inactive), Silencing mechanism involves spreading of heterochromatin and can spread into euchromatin regions, once region is established it is then stably inherited.

        - Genomic imprinting- Primarily driven by DNA methylations, during gamete formation specific DNA regions are methylated; its sex specific so eggs and sperm have paternal and maternal methylation patterns respectively, after fert parental meth is maintained in somatic cells of the offspring, methylated allele silenced, unmethylated remain active, in germline of next gene, imprint marks are erased.

        - X-chromosome inactivation- Process that occurs in female mammals to equalize the gene dosage of X-linked gene between males and females. Each female somatic cell, one of the two X chromosomes is randomly and permantly silenced, resultes in female being genetic mosaic

- Detail the following PEV mechanisms

        - Chromosomal rearrangement- Moving a gene a from an active location to adjacent inactive location

        - Spreading of heterochromatin- Heterochromatin regions can spread into euchromatin regions

        - Epigenetic inheritance- Once a heterochromatin region is established, it is stably  by all of its daughter cells