Components of a Cell

DNA must be protected (it is fragile) and accessed in a controlled way. the cell isolates DNA in a dedicated compartment → —

the nucleus

Key consequences:

• The nucleus is the main site of —-.

• It is also where RNA is made (—-), since RNA is a working copy of DNA instructions.

• DNA replication

• transcription

Only small amounts of DNA exist outside the nucleus (—)

mitochondria

Once instructions exist, the cell needs a place to do the work. The working space → —

Cytoplasm and Cytosol

Everything outside the nucleus is the —

cytoplasm.

The fluid part of the cytoplasm is the —

cytosol.

Why the cytosol matters:

• It is where:

  • Most proteins are —

  • Most proteins are degraded

  • Most intermediary metabolism happens (breaking down things into AA, nucleotides, etc)

• synthesized

Instructions (RNA) are useless unless they are turned into —

functional molecules (proteins)

— read RNA and build proteins.

Ribosomes

Why the Endoplasmic Reticulum (ER) exists

Problem:
Some proteins must:

• —-

• Be secreted (AKA be in vesicles)

Solution:
Make them directly into a membrane system → the ER.

why does this solve the problem?

because we need to get the vesicles from somewhere and the ER buds off the vesicle to enclose the proteins that need to be secreted

Key first-principle insight:
Proteins enter the ER while they are being synthesized, not after.

what is meant by this?

when we talk about these secreted proteins, they first start being made in the cytoplasm like everybody else, but then they finish translation in the ER

why do we need some free ribosomes and some attached to the ER

• some will be fully translated in the cytoplasm so we need free ribosomes

• some will start in cytoplasm and then finish in ER, so we need ribosomes there too to finish the job

smooth er is mostly for

calcium storage

Making molecules isn’t enough, they must be modified and delivered correctly. Sorting and shipping → —

The Golgi Apparatus

The ER sends proteins to the —

Golgi apparatus

What the Golgi does:

1. Receives proteins from the ER

2. —- them

3. Sorts and dispatches them

1. Chemically modifies

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Only proteins that enter the ER can ever go to the Golgi.
Proteins made in the cytosol never go to the Golgi.

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the only way into the Golgi is:
👉 inside a vesicle

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All proteins start on ribosomes in the cytosol.

t

At the beginning, every ribosome is:

• Free in the cytosol

• Making the first few amino acids of a new protein

if this protein needs to be secreted-HOW DOES IT KNOW IT NEEDS TO GO TO THE ER?

A protein contains address information in its amino acid sequence.

Think of this as a shipping label embedded in the protein itself.

Only proteins with an ER signal sequence go to the ER.

for a secreted protein:

As the protein is being synthesized, an— emerges

ER signal peptide

the ER signal peptide is recognized by the —

Signal Recognition Particle (SRP) (this is a whole other separate player just floating aroudn scanning for the signal, it is not part of the ribosome)

— pauses translation

Signal Recognition Particle

after SRP gets involved and pauses translation, what happens next?

The ribosome is moved to the ER membrane

This secreted translation pathway exists because:

• Secreted proteins MUST be put in—

• Vesicles bud only from the endomembrane system (ER → Golgi → membrane)

• vesicles

• By endomembrane system, we mean vesicles are capable from budding from those organelles, therefore, if we need to get it to the Golgi, it MUST have gotten a vesicle from the ER

what does “secreted protein” actually mean?

A secreted protein is a protein whose final functional location is outside the cell.

(blood, extracellular matrix, insulin, etc)

Why can’t the cell just make secreted proteins in the cytosol and let it drift out?

Problem #1: The plasma membrane is a barrier

• Proteins are large and polar

• They cannot cross lipid bilayers by diffusion

So if you make insulin in the cytosol:

• It is trapped inside the cell

• There is no hole it can just walk through

The only safe way to move a protein across a membrane is:

—-

Keep it enclosed by a membrane (vesicle) the entire time

a vesicle is

A small bubble of membrane

esicles cannot form out of thin air.

They must:

• Bud off from—

• an existing membrane

First-principle problem

Cells need a constant supply of ATP to drive function

Solution

Use a specialized compartment to:

• Maintain proton gradients

• Run redox reactions efficiently

this specialized compartment is —

the mitochondria organelle

First-principle problem

Cells must:

• Break down worn-out organelles

• Digest material taken up from outside

• Do this without destroying themselves

Solution:

Create a sealed compartment full of digestive enzymes:
→ This is the Lysosome

Why lysosomes MUST be vesicular

• —- are dangerous

• If released into cytosol → cell death

• Must be isolated behind membranes

• Digestive enzymes

The organelle itself is not “built” like a machine — it is a space.

• Nucleus = membrane + contents

• Mitochondrion = membranes + contents

• Lysosome = membrane + contents

its the contents that are proteins that perform functions, the membrane is just to create a space, a compartment

ok

so basically, the lysosome itself, it just a membrane

it contains lysosomal enzymes (digestive) which were synthesized in the

a) cytoplasm

b) ER

ER

they need to travel somewhere specific so they get “shipped” to the lysosome by a vesicle system and then put into another vesicle (the lysosome itself)

First-principle problem

Material taken into the cell (endocytosis) must be:

• Sorted

• Recycled

• Or sent for destruction

Solution

—

Create intermediate compartments:
→ Endosomes

are endosomes basically vesicles?

yes, again it’s literally just a membrane bubble that encloses a space

What endosomes do

• Receive vesicles from the —

• Sort contents:

  • Some back to the membrane

  • Some onward to —-

• plasma membrane

• lysosomes (destruction)

You don’t want everything you ingest to be immediately destroyed, this is why — are helpful

endosomes

First-principle problem

Some reactions:

• Produce harmful byproducts (e.g. hydrogen peroxide)

• Should not occur in cytosol

Solution

Isolate these reactions in small compartments:
→ This is what Peroxisomes are

difference between lysosome and peroxisome

lysosome is more for breaking things down, garbage can, more general

peroxisomes have the specific function of detoxifying harmful substances, they use a different system to break things down and a lot of toxic stuff goes on in there

t or f'

If every cell had the same organelle proportions, most cells would be inefficient

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Why the Golgi sits near the nucleus

• The Golgi receives proteins from the ER

• ER is continuous with the —

• Shorter distances = faster trafficking

• nuclear envelope

This minimizes transport time- the point of this is to say that the position of organelles inside the cell matters too, it’s not random

The precursors of the first eukaryotic cells are thought to have been relatively simple cells that —like most bacterial and archaeal cells—have a—-

plasma membrane but no internal membranes

That one membrane did everything:

• Pumped ions

• Made ATP

• Secreted proteins

• Synthesized lipids

eukaryotic cells are a lot bigger, but why couldn’t this one-membrane-system just scale up?

So a large cell with only one membrane has a crisis:

• Too little membrane area (surface area) to support the volume

• Too many membrane-based tasks

Evolution’s solution: -

internal membranes

the only way to:

• Increase membrane area

• Without increasing cell size

is to: fold the plasma membrane inward (invagination) just like brain gyri, cristae, it’s to increase surface area

Why are mitochondria so different from every other membrane-enclosed organelle?

Because they did not evolve from membrane invagination.
They evolved from engulfed bacteria

So far you’ve learned about organelles that came from:

• Plasma membrane folding inward
  (ER, Golgi, lysosomes, endosomes, etc.)

Now mitochondria belong to a completely different category.

okay

Imagine an ancient cell:

• It had no internal membranes

• It engulfed a bacterium

• But instead of digesting it, the two formed a —

• mutually beneficial relationship

because

The bacterium:

• Was good at generating ATP

Over evolutionary time:

• The bacterium became a permanent resident

• It turned into what we now call a —

This is endosymbiosis.

• mitochondrion

Why mitochondria have their own genome

If mitochondria came from bacteria, then originally they:

• Had their own —

• Had their own ribosomes

• Controlled their own replication

• DNA

During engulfment:

• The bacterium already had its own plasma membrane

• The host cell wrapped a membrane around it

So mitochondria ended up with:

1. Inner membrane → —

2. Outer membrane → —

1. Inner membrane → original bacterial plasma membrane

2. Outer membrane → host-derived engulfing membrane

The mitochondrial matrix = former —

bacterial cytoplasm

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• No vesicles bud to mitochondria

• No vesicles bud from mitochondria

• Their membranes are not part of the endomembrane system

t

This isolation is not a design choice — it’s historical inevitability.

problem:

DNA needs protection and regulation, but gene products must reach the cytosol efficiently.

Solution :

—

• Enclose DNA in a nucleus

• Add nuclear pore complexes instead of vesicles

No membrane fusion or vesicles involved

for things that don’t go through pores or don’t go in vesicles, what is the only other option?

The transported protein molecule usually must unfold to snake through the translocator

For a cell to stay alive and function, it must be able to:

1. Hold a shape (not collapse like a water balloon)

2. Withstand forces (pushing, pulling, squeezing)

3. Organize its inside (proteins, organelles can’t float randomly)

4. Change shape (during growth, division, movement)

5. Move things internally (vesicles, chromosomes, organelles)

6. Move itself (migration, tissue formation, immune response)

What I the solution?

the cytoskeleton: which is a flexible but rigid network of proteins

Why not one solid structure like bones?

Because cells need:

• Strength and flexibility

• Stability and rapid change

what is the name of the microfilament?

actin

what is actin used for?

cell shape and movement (crawling, contraction)

One important property of actin filaments?

Rapidly assembling/disassembling

what is the name of the “microtubules”?

tubulin

what are the three structural elements of cell (proper names)

1. actin (microfilament)

2. tubulin (microtubule)

3. Intermediate filaments

its hard to come up with one borad term for what microtubules are used for but they’re really Important in (2)

• forming the mitotic spindle to pull apart chromosome sin mitosis and meiosis

• kinesin and dyenin are motor proteins of microtubules

when you see intermeditate filament, think —

Provides Mechanical strength

(the thing in nails and hair)

Keratin is an intermediate filament

Structures actin builds:

Dynamic protrusions

• —-: broad, sheet-like extensions for crawling

• —: thin spikes for sensing the environment

Highly ordered bundles

• — (inner ear): rigid actin bundles tilt as rods → convert sound vibrations into signals

• — (intestine): actin-supported projections increase surface area for absorption

Important structure in mitosis and meiosis:

• Lamellipodia

• Filopodia

• Stereocilia

• Microvilli

CONTRACTILE RING!

also acts in muscle contraction

Structures microtubules build:

• -

• -

• -

-Mitotic spindle

-cilia

-flagella

-axons

in microfilaments, the subunit is:

in microtubules, the subunits are:

actin

alpha and beta tubulin (dimer)→ together called tubulin

Cytoskeleton and Membrane Trafficking, why is this Important?

remember everything we just learnt about vesicle movement? Well, vesicles can’t just move on their own, they are carried on highways by microtubules and the cytoskeleton with motor proteins like kinesin, dyenin, myosin

not just vesicles, but also organelles like mitochondria, endosomes etc

Cytoskeleton and Phagocytosis

• Actin filaments rapidly polymerize at the site of the bacteria to push the membrane around it.

• This forms pseudopods that wrap the particle. (ARMS)

principle behind dyes

• Binds preferentially based on charge, hydrophobicity, or structure

hematoxylin binds:

negatively charged molecules (DNA)

what problem is in situ hybridization trying to solve?

In this tissue, which cells are expressing gene X, and how much?

it’s harder to get proteins so, the soltuion:

instead of detecting protein → detect mRNA directly

how is in situ hybriziation done?

• fix cells: this preserves mRNA

• section tissue

• diffuse your fluorescent probe inside (probe is just a complimentary binding thing)

• probe binds to mRNA (regardless that the cell is dead, it can base pair)

• use a fluorescent microscope to visualize localization of expression (can quantify with softwares)

• qPCR / RNA-seq → quantitative claims

• FISH / ISH → spatial, cell-type, and — claims

• localization

FISH is excellent for:

1. Which cells express gene X?

2. Where in the tissue is it expressed?

3. Is expression uniform or heterogeneous?

4. Is transcription active right now? (nuclear spots)

5. Does expression change in specific cell types?

okay

• mRNA ≠ protein (always)

• Western blots validate functional output

so basically, if they do something related to mRNA in the study, say that mRNA is just a proxy for protein so they should validate with protein study like —-

PROTEOMICS!

mass spectrometry is basically the RNA-seq equivalent

distinction between when you would use western blotting and immunohistochemistry

Western blot → how much protein (it’s extracted from cell, you get visual band output)

IHC → where protein is IN the cell (you preserve the netire cell, so you can see the relative amount as well as the spatial localization)

First principle: cells are dynamic, not static

Earlier techniques (fixation, staining, FISH, IHC):

• Kill the cell

• Give a snapshot

Issue?

But cells:

• Move proteins

• Assemble/disassemble structures

we want a ay to watch the dynamics in real time

solution?

genetically encoded fluorescence with GFP (no dyes, no antibodies needed)

Whenever the gene is transcribed → GFP is made

GFP fluorescence = gene expression (proxy for)

1. GFP as a reporter of gene expression -What question this answers

“When and where is this gene turned on?”

⚠ Important limitation:

• GFP reports —, not native protein behavior

• transcription only

FRET answers:

Are these two proteins physically interacting right now?

What FRET is used for

• —

• Signaling cascades

• Protein complex assembly

• Receptor–ligand interactions

FRAP measures:

Protein mobility and kinetics/turnover movement

FRAp stands for

Fluorescence Recovery After Photobleaching