lala added ch 16

Negative Numbers ( -100, -1000)

Positions Upstream of the start site

diffeential gene regulation

Cells of different tissues express different sets of genes

Why does transcription initiation in eukaryotes require numberous proteins and allosteric changes?

Assembly of the transcription complex requires multiple proteins (activators, coactivators, RNA Pol II, GTFs) and allosteric changes that allow them to properly bind each other and DNA, ensuring efficient transcription initiation

transcription initiation complex

exists to assemble proteins at the promoter to position RNA polymerase and unwind DNA

Enhancer

DNA sequence that increases transcription when bound by activators.

Where are enhancers found?

Upstream, downstream, or within introns-- or far from gene

"Action at a Distance"

Enhancers affect transcription from far away via DNA looping

"Upstream"

Region before the transcription start site.

"Downstream"

Region after the transcription start site.

+1

transcription start site

Cell Differentiation

Cells becoming specialized for structure & function; occurs during embryonic development and can continue into adulthood

positive feedback loop

Some Transient Signal turns on expression of protein A, making transcription factor activate its own gene. Once its on it continues to be produces even after initial signal is gone. This memory allows cell to remember its identity and maintain differentiated state over time

stem cell

unspecialized cell that can develop into a specialized cell under the right conditions

RNA Polymerase

transcription enzyme

All genes...

have a series of enhancer sequences with specific regulatory proteins

Similarities of prokaryotic and eukaryotic gene regulation

Both regulate gene expression with DNA-Binding proteins & respond to environmental signals

Prokayotic gene regulation

operons, transcription and translation are couples, simpler regulation

Eukaryotic gene regulation

Complex regulation, changes in chromatin structure, transcription occurs in nucleus, many levels of control

RNA Pol I

transcribes rRNA

RNA Poly II

transcribes mRNA and some snRNA

RNA Poly III

transcribes tRNA and 55 rRNa

general transcription factors (GTFs)

Correctly position polymerase at/around start site and help intiate transcription. analogous to Bacteria Sigma Factor

TFIID/TBP

bind to TATA box using TBP, bending DNA and allowing other proteins access

TFIIH

releases Pol II by phosphorylation of the C-Terminal tail of RNA Pol II. Leads to start of transcription

How do activators stimulate transcription through direct association

Activators directly bind RNA Pol II/GTFs to increase transcription

How do repressors inhibit transcroption through direct association

By binding/interfering with transcription machinery

How do activators and repressors regulate transcription by chromatin modification

Activators: reruit chromatin-modifying enzymes to open DNA

Repressors: Promote Chromatin condensation, making DNA less accessible to transcription machinery

How do HATs affect chromatin structure

add acetyl groups to histones that loosen the chromatin --> increasing transcription

How do HDACs affect chromatin structure?

remove acetyl groups, tightening chromatin --> decreasing transcription

How does the glucocorticoid receptor regulate genes

A ligand-activated transcroption factor that binds to glucocortid hormones, enters nucleus, and regulates gene ecpression by binding specific DNA sequences

Why is a skin cell different from a muscle cell?

They're diff because they express different genes, allowing them to produce different proteins and giving each cell a unique structure & function

If DNA is a molecule with Partial and Full charges

then it can form non-covalent bonds w/ proteins with polar and charged amino acids

What is the significance of the phosphorylated tail of RNA Pol II

- start of elongation

- recruits RNA processing proteins (capping, splicing, polyadenylations)

- Coordinates initiation --> elongation transition, integrating transcription with RNA maturation

How do cells control gene expression in a temporal manner?

By controlling the expression of regulatory factors allowing genes to be ecpressed only when needed

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Nuclear Import

1. Protein contains a nuclear

localization signal (NLS) (basic)

2. Binds to a specific receptor protein

(importin) via the NLS

3. Importin associates with nuclear

pore

4. GTP-mediated import

5. Complex released inside nucleus

Nuclear Export

1. Protein contains nuclear export signal

(NES) (leucine-rich)

2. Protein binds to specific receptor

protein (CRM-1) via the NES

3. CRM-1 associates with nuclear pore

4. GTP-mediated export

5. Complex is released outside nucleus

6. Protein may be associated with RNA.

What are sorting signals?

Amino acid sequence that specifies where a protein is sent.

What are sorting signals made of?

Specific, short amino acid sequences (5-60 residues) within proteins that act as molecular "zip codes," determining their proper destination within or outside the cells

What happens when a signal sequence is swapped btwn proteins that function in a different subcellular compartment?

the protein will be trafficked to the wrong location. Proteins cannot function in the wrong environment --> leads to degradation or incorrect cellular function

What do sorting signals do? Where are they found?

Sorting signals are at the N-Terminus, hydrophobic and part of the primary structure of the protein. Used to sort proteins, insert protein into organelle/membrane

Where do chloroplast and mitochondrial proteins originate from?

Nuclear Genome & Organelle's Genome. endosymbiosis

Where are ribsosomes made?

Nucleolus

What are free ribosomes

Riobosomes in cytosol making proteins for cytosol, nucleus, mitochondria, peroxisomes

What are bound ribosomes?

Riobosomes attached to rough ER making secreted, membrane, or lyossomal proteins

Do ribosomes start as free or bound?

Free

What makes a ribosome become bound?

Signal sequence and SRP (signal recognition particle) targeting to ER

What happens to ribosomes after translation?

They detach and become free again

Difference bewtween free and bound ribosomes?

Depends on protein being made

The N-Terminus is

nonpolar, facing the cytosol

Proteins are sorted to locations beyond the ER within

transport vesicles

Secetory Pathway (exocytosis

-for export

- deliveries lipid/protein to plasma membrane

Endocytic Pathway

- for import

How do vesicles form?

Budding driven by assembly of a protein coat

How is vesicle formation controlled?

Different membranes have their own protein coats

Clathrin

protein that coats the plasma membrane's inward-facing surface and assists in forming specialized structures, like coated pits, for phagocytosis

coated pit becomes

a coated vesicle

Adaptin

binds clathrin and cargo receptors (an adaptor protein)

Endocytosis at Plasma Membrane

1. Binding of cargo to cargo receptors

2. Lateral movement into a coated pit

3. Binding of adaptin to receptor

4. Binding of CLATHRIN to adaptin

5. BUDDING of coated pit

6. Vesicle is pinched off by dynamin

(GTP)

7. Uncoating

8. Onward to target

Enables endocytosis to occur

What needs to happen in budding/fusion?

• Each vesicle must take ONLY those

proteins destined for target.

• Each vesicle must fuse with

CORRECT membrane.

• Lipid MAKE-UP of target must be

conserved.

• AMOUNT of lipid in target must be

conserved.

The cytosolic face

remains facing the cytosol

How does a vesicle know where to go?

Specific protein in its membrane (aka v-Snare)

Target membrane is the receptor (T-Snare)

How does a vesicle reach its destination?

Uses micotubules as tracks w/ motor proteins using ATP hydrolysis (conformational change)

oligasaccharides

short chain of sugar residues/ often lipid linked to protein

Glycosylation

enable cell to cell communication using sugars on the cell suface

Protein folding in the cell is facilitated by

chaperon proteins

Chaperon proteins

- retain misfolded proteins in ER then degrades in a lysosome

Proteins move through golgi via

vesicles

cisternae

flattened stacked membrane folds that make up the golgi apparatus

Cisface

receiving side of golgi apparatus; recieves vesicles

Transface

shipping side of golgi apparatus; buds & sends off vesicles

Golgi cisternae...

further modify proteins by adding or removing sugars to the oligosaccharides

Constitutive pathway (think constant)

- operates continually

- supplies lipids, membrane proteins, and soluble proteins to cell membrane

Regulated pathway-- when the time is right..

- operates in SPECIALIZED SECRETORY CELLS (e.g glands)

-secretory proteins are stored in specialized secretory vesicles (hormones, digestive enzymes, etc)

pinocytosis

- cellular drinking

- generalized

Endocytic pathway

form plasma membrane to a lysosome

-> pinocytosis

->phagocytosis

Phagocytosis

- "cell eating"

- large

-specific cells do dis

Endocytosis

continual intake of vesicles and recycling of surface receptors

-> non receptor mediated

-> receptor-mediated (e.g protein coating)

Endosome

destination of vesicles

function of the endosome

- Allows cargo to come apart from receptor originally bound to

- receptor is recycled and sorted back to cell surface

-control for endocytosis

why does cargo come off receptor?

The endosome is slightly acidic.

One function of the golgi

- control for exocytosis

High cholesterol is from

a mutation in LDL receptor

Receptors can be

1. recycled to the same memrbane

2. degraded in the lysosome

3. moved to another suface with ligand

Lysosomes

Sites of intracellular digestion of extracellular compounds

How do enzymes get to the lysosome?

theyre made by ribosomes on the ER surface, sorted to the golgi, then the endosome, then finally the lysosome

Why do enzymes in the lysosome not destroy the ER

The environment of the lysosome has a pH of 5, snapping the acidic hydrolytic enzymes into an active conformation —> enzymes are active at a low pH but a lysosome is acidic.

Why is the interior of the lysosome acidic

proton pumps in the membrane of the lysosome; it is an ATP driven pump

What is a phospholipid

It is a lipid that has a phosphate group attached to the glycerol and only two fatty acid chains.

What is the phospholipid head composed of?

Glycerol, phosphate, and a chemical group (choline, ethanolamine, serine and inositol). hydrophobic and hydrophilic regions make phospholipids amphipathic

What keeps the closed compartments made of phospholipids

Phospholipids naturally form bilayers and sealed compartments in water in ball formations. Thermodynamically this is the most favorable conformation of phospholipids

What part(s) of the membrane is fluid?

The phospholipid bilayer

What motion occurs in the membrane?

-Lateral diffusion

-rotation

-flip-flopping (rare, fixed by flippases)

Why is the membrane fluid?

Fatty-acid tails move laterally, where fluidity originates

How does the saturation-level of fatty acid tails affect membrane fluidity

"Saturated" --> max # of Hydrogens, no kinks in tail. Better packing therefore the lipid is less fluid

"Unsaturated" --> less than the max # of H which causes kinks due to double bonding; kinks mean less packing which means lipid is more fluid

How does the presence of cholesterol affect membrane fluidity?

High temps --> reduces fluidity by restricting phospholipid movement

Low temps --> increases fluidity by preventing tight packing & freezing.

What is cholesterol? Where is it located?

Cholesterol is embedded in the membrane (20% of lipid) and decreases permeability by restricting lipid movement which prevents crystallization

Located in the top half of phospholipid tails

Why is fluidity important?

1) membrane permeability

2) protein functioning (e.g protein-protein interactions)

3) redistribution of lipids/proteins (i.e synthesis, fusion, cell division)

How are membrane proteins associated with membranes?

- Transmembrane (integral)

- Membrane associated (integral)

- Lipid linked (integral)

- Protein-protein (peripheral)