Regulation of Gene Expression and Protein Metabolism

______

under constitutive expression

“always on”

constantly expressed at stable levels in approximately all cells

Housekeeping gene

______

Levels of the gene product rise and fall with the needs of the organism.

Turned “on” or “off”

Such genes are inducible.

able to be turned on

Such genes are also repressible.

able to be turned off

Regulated gene

_____

They reduce RNA Pol-promoter interactions or block the polymerase (RNA Pol)

Bind to operator sequences on DNA

usually near a promoter in bacteria but further away in many eukaryotes

Effectors can bind to repressor and induce a conformational change.

change may increase or decrease repressor’s affinity for the operator and thus may increase or decrease transcription

Repressors

______ involves repressors.

(a) Repressor binds to the operator in the absence of the molecular signal; the external signal causes dissociation of the repressor to permit transcription.

(b) Repressor binds in the presence of the signal; the repressor dissociates and transcription ensues when the signal is removed. Positive regulation is mediated by gene activators.

Negative regulation

_____ involves activators.

Enhance activity of RNA polymerase

Activator may remain bound until a molecule signals dissociation  stop transcription

Alternatively, the activator may only bind when signaled as part of the assembly of the transcription complex

Positive regulation

______ Allows Eukaryotic Enhancers to Be Far from Promoters

Eukaryotic activators and repressors often bind sites thousands of base pairs from the promoters they regulate.

DNA looping, often facilitated by architectural regulators, brings the sites together.

The interaction between activators and RNA polymerase is often mediated by coactivators, as shown.

Repression is sometimes mediated by repressors that bind to activators, thereby preventing the activating interaction with RNA polymerase.

DNA Looping

______: A few protein arrangements are used in thousands of different regulatory proteins

helix-turn-helix

zinc finger

leucine zipper

Motifs

The _______ Is Common in DNA-Binding Proteins\

• ~ 20 amino acids

  one a helix for recognition for DNA (red), then b turn, then another a helix

  sequence-specific binding due to specific contacts between the recognition helix and the major groove

  Four DNA-binding helix-turn-helix motifs (gray) in the Lac repressor

Helix-Turn-Helix Motif

~30 amino acids

“Finger” portion is a peptide loop cross-linked by Zn2+

Zn2+ usually coordinated by four Cys, or two Cys and two His

Interact with DNA or RNA

Binding is weak, so several zinc fingers often act in tandem.

Binding can range from sequence specific to random.

The Zinc Finger Motif

Dimer of two amphipathic a helices plus a DNA-binding domain

Each helix is hydrophobic on one side and hydrophilic on the other.

The hydrophobic side is the contact between the two monomers.

Approximately every 7th residue in helices is Leu.

Helices form a coiled coil.

The DNA-binding domain has basic residues (Lys, Arg) to interact with polyanionic DNA.

The Leucine Zipper Motif

Features of Eukaryotic Gene Regulation:

Access of eukaryotic promoters to RNA polymerase is hindered by chromatin structure.

this requires ______

Positive regulation mechanisms predominate and are required for even a basal level of gene expression.

Eukaryotic gene expression requires a complicated set of proteins.

remodeling chromatin

________ = less-condensed chromatin, distinguished from transcriptionally inactive heterochromatin

Chromatin remodeling of transcriptionally active genes:

nucleosomes repositioned

histone variants

covalent modifications to nucleosomes

Euchromatin

Histone Modification Alters Transcription

Covalent modification of histones allows recruitment of enzymes and transcription factors.

____ of Lys-4 and Lys-36 at histone3 (H3) and Arg of H3 and H4:

results in transcriptional activation

recruits ______ that then acetylate a particular Lys

reversed by ________ that make chromatin inactive

Acetylation of Lys results in decreased affinity of histone for DNA.

Methylation, histone acetyltransferases (HATs), histone deacetylases (HDACs)

Activators can influence transcription at promoters thousands of bp away.

Looping can be facilitated by _______

Co-activators may mediate binding by binding to both activator and RNA polymerase.

DNA Looping Allows Eukaryotic Enhancers to Be Far from Promoters

architectural regulator proteins

_____

Requires 5 components

Transcription activators (enhancers)

proteins that bind to upstream activator sequences (UASs)

Architectural regulators to facilitate DNA looping

Chromatin modification/ remodeling proteins

Coactivators

act indirectly (with other proteins, not with DNA)

Basal (general) transcription factors

RNA Polymerase II

Coactivators Assist RNA Polymerase

_____ binds to carboxyl-terminal domain (CTD) of RNA Pol II

required for transcription at many promoters

later provides assembly surface for other complexes

_______ is first component of preinitiation complex (PIC) at the typical TATA box of a promoter

Mediator complex, TATA-binding protein (TBP)

Four Mechanisms of Translation Regulation

Phosphorylation of translation initiation factors

Translational repressors (typically bind to 3’ UTR)

Disruption of eIF4E and eIF4G interactions

RNA-mediated regulation (gene silencing)

______ silence genes by binding to mRNAs

can prevent transcription of the mRNA by cleaving it (via _______) or by blocking it

Some miRNAs are made briefly during development

small temporal RNAs (stRNAs).

Micro-RNAs (miRNAs), endonucleases Drosha or Dicer

Any dsRNA that corresponds to an mRNA and is introduced into a cell will be cleaved by Dicer into short segments called small interfering RNAs (siRNAs).

These will bind to the mRNA to silence its translation.

The process is called _____

RNA interference