FINAL EXAM 4 MCB Questions

What does the cell microenvironment include?

Local environment, signals, and behavior; two regulatory components: physical structure/mechanical and signaling (auto, endo, para).

What are the two regulatory components of the cell microenvironment?

Physical structure/mechanical and signaling (autocrine, endocrine, paracrine).

Give a healthy example of microenvironment influence in the mammary gland.

Pregnancy vs puberty changes, with timing/milk production.

What happens to HIF1 and O2 in angiogenesis?

↑HIF1 occurs with ↓O₂; ↓HIF1 occurs with ↑O₂.

Why is the TME important?

It is important in cancer biology; it consists of cellular components, secretory factors, and the ECM.

Is the TME hypoxic or oxygen-rich, and what does that promote?

Hypoxic; it promotes HIF1.

Why are fibroblasts key in tumors?

They are involved in tumor initiation/progression, auto/paracrine signaling, and facilitating angiogenesis/cancer (CAFs); they also act in wound healing.

What enzymes are associated with fibroblasts in the TME?

Matrix metalloproteinases (MMPs).

What does CAF stand for?

Cancer-associated fibroblast.

What key growth factor is associated with CAFs?

Transforming growth factor β (TGFβ).

List the cancer-promoting effects of CAFs.

Cancer proliferation, angiogenesis, invasion, metastasis, and modulation of inflammation.

What is TGFβ's role in tissue injury?

It maintains homeostasis in tissue injury.

Describe TGFβ as a sensor.

It is highly oxidized and acts as an extracellular sensor of oxidative stress.

Which signaling pathway does TGFβ use?

The SMAD-dependent signaling pathway.

What are the targets of TGFβ signaling?

Immunosuppression, inflammation, and EMT/invasion.

What are the cellular responses of TGFβ signaling in the nucleus?

Tumor suppressor activity, environment modification (fibrosis), and phenotypic plasticity.

Which SMADs are phosphorylated by the TGFβ receptor and which one complexes with them?

SMAD2/3 are phosphorylated and complex with SMAD4.

What other pathway can the TGFβ receptor activate, per the diagram?

Ras → Erk.

List therapies targeting CAFs/fibroblasts.

Target activated fibroblasts, fibroblast senescence, inflammation inhibitors, target fibroblast activation protein (FAP), and antioxidants against ROS (ascorbate).

What does aging of fibroblasts lead to?

Pancreatic cancer.

What combination targets the TME to halt progression?

ALOX12 + 12-(S)-HETE.

List the characteristics of stem cells.

Unspecialized, no shape/function, divide asymmetrically (fate decision needed).

List the characteristics of differentiated cells.

Specialized function, cannot be altered, symmetric division; some can still divide into progenitor cells.

What dictates a cell's identity?

The pattern of gene expression (structure dictates function).

What are housekeeping proteins/RNAs?

Proteins/RNAs present in all cells, with sequence ID conserved between species.

What are biomarkers?

Proteins unique to a cell type that are used to identify the cell type.

Describe Mechanism A of daughter cell fate choice.

Asymmetric division: internal signal, fate fixed in each lineage; used in DEVELOPMENT; involves gene expression regulation and protein sorting & trafficking.

Describe Mechanism B of daughter cell fate choice.

Independent choice: the cellular microenvironment determines daughter cell ID, random choice; used in REGENERATION/REPAIR via secreted molecules, physical contact, and chemical environment.

Which process uses asymmetric division, development or regeneration/repair?

Development.

Define unipotent.

One cell type (self renewal).

Define multipotent.

Some cell types (adult stem cells).

Define pluripotent.

Most cell types (engineered stem cells).

Define totipotent.

All cell types (zygote).

Where are neural stem cells found and what do they become?

Adult brain (SGZ, SVZ); neurons, oligodendrocytes, and astrocytes.

What are NSC biomarkers?

PCNA & MCM2, Nestin, Sox2/SRY.

What are the sources of HSCs?

Bone marrow and umbilical blood.

What do HSCs differentiate into?

RBCs, WBCs, and platelets.

What are HSC biomarkers?

CD proteins.

What are the sources of MSCs?

Bone marrow, dental pulp, umbilical blood, adipose tissue, placenta.

What do MSCs differentiate into?

Bone, cartilage, muscle, fat storage cells, and CT cells.

What are MSC biomarkers?

Too diverse.

What determines the ID of epithelial stem cells?

The cellular niche (microenvironment).

What happens to the cell cycle potential of EPSCs with age?

It is infinite but decreases with age.

Where are EPSCs found?

Skin appendages, intestinal lining, mammary tissue.

What are EPSC biomarkers?

Integrins, E-cadherins, keratins.

How do heart stem cells behave?

They are slow and decline with age.

What is notable about lung stem cells?

There are rare varieties, and disease states alter them.

Describe Option 1 for engineering pluripotent stem cells.

Embryonic stem cells (ESCs): harvest the inner cell mass of the blastocyst, which destroys potential life.

Describe Option 2 for engineering pluripotent stem cells.

Somatic cell nuclear transfer (SCNT): nucleus of a skin cell is placed into an oocyte (patient specific).

Describe Option 3 for engineering pluripotent stem cells.

Induced pluripotent stem cells (iPSCs): reprogram somatic cells (patient specific).

Which three genes induce the stem cell state?

Oct4, Sox2, and KLF4.

Which gene is linked to cell cycle control in stem cells?

c-Myc.

Which gene maintains the stem cell state?

Nanog.

Describe Oct4.

Expressed in early embryos; a transcription factor that activates its own expression.

Describe Sox2.

Complexes with Oct4 and binds DNA.

Describe KLF4.

A zinc finger DNA binding protein; a transcription factor.

Describe c-Myc.

Mutated in cancer; a cell cycle promoting factor; a helix-loop-helix + leucine zipper transcription factor.

What does Nanog do?

Maintains the ESC phenotype; activates stem cell genes and represses differentiation-inducing genes.

How do Hox and Nanog relate?

They negatively regulate each other; Hox triggers differentiation.

List the important signaling pathways in stem cell vs differentiated cell decisions.

TGF-β (SMAD2/3), FGFR (Ras/Raf/MAPK/Akt), BMP (SMAD1/5/9), Notch, and Wnt.

What is BMP's effect on Nanog?

BMP (SMAD1/5/9) is a Nanog inhibitor.

What type of signaling is Notch?

Paracrine signaling.

What does Wnt do?

The protein interacts with GPCR and integrates signals from all pathways.

Why engineer stem cells?

Organoids, study basic biology, drug safety testing, tissue for transplants, preventing/treating genetic disease, and gene therapy to correct genetic disease.

What is the approved stem cell therapy mentioned?

HSCs derived from umbilical cord blood.

Define apoptosis.

Routine programmed cell death; it is clean, and cells shrink and condense.

List the phenotype of apoptosis.

Shrinkage of cell/nucleus volume, loss of adhesion, blebs on surface, DNA fragmentation, cytoskeleton collapse, nuclear envelope disassembly, and rapid engulfment by phagocytosis.

What are the characteristics of apoptosis?

DNA fragments (endonuclease cleaves), cleavages in linker regions of nucleosomes, and cytochrome C released from mitochondria.

What are caspases?

Cysteine aspartyl specific proteases with a cysteine in the active site; apoptosis is a proteolytic cascade and their activation is the key event.

What are procaspases?

Inactive precursors/monomers activated by protease cleavage; once cleaved they form large + small subunits (heterodimer).

What activates procaspases?

Other caspases, and an apoptotic signal → adaptor protein complex.

What are the two major classes of caspases?

Initiator (caspase 8 & 9) and executioner (caspase 3).

What do initiator caspases do?

Initiate apoptosis (caspase 8 & 9).

What do executioner caspases do?

Destroy targets: cleave downstream proteins, cleave inactive endonuclease, target the cytoskeleton, and attack cell adhesion proteins to roll the cell into a ball.

What are the two apoptosis pathways?

Extrinsic and intrinsic.

Is the extrinsic pathway mitochondria dependent?

No, it is mitochondria independent.

What triggers the extrinsic pathway?

Extracellular signals binding death receptors.

What structure do death receptors have?

They are homotrimers with 3 domains: extracellular, single transmembrane, and intracellular death.

Step 1 of the extrinsic pathway?

Fas ligand binds the Fas death receptor (homotrimer).

Step 2 of the extrinsic pathway?

FADD adaptor + initiator procaspase-8 are recruited with death domain (DED).

Step 3 of the extrinsic pathway?

FADD + procaspase-8 form trimers, bringing death domains together → DISC.

Step 4 of the extrinsic pathway?

Active initiator caspase 8/10, then active downstream executioner caspase-3 & caspase-7.

Step 5 of the extrinsic pathway?

Apoptosis.

What does DISC stand for in the extrinsic pathway notes, and how is it formed?

DISC is formed when FADD and procaspase-8 form trimers that bring death domains together.

How do decoy receptors inhibit the extrinsic pathway?

They have no DED domain and act as sponges; they bind domain → no apoptosis.

What is FLIP?

A competitive inhibitor against procaspase-8/10 (a sponge) that prevents apoptosis.

What is the intrinsic pathway?

Activation of apoptosis from inside the cell in response to injury/DNA damage.

What is the key event of the intrinsic pathway?

Translocation of cytochrome C from the intermembrane space of mitochondria into the cytosol.

What does cytochrome C bind in the cytosol?

The procaspase-activating adaptor protein Apaf1.

What forms when cytochrome C binds Apaf1?

The apoptosome, which activates initiator caspase-9.

What does caspase-9 activate in the intrinsic pathway?

Executioner caspase-3 → apoptosis.

What family of proteins controls apoptosis?

The Bcl2 family.

What does Bcl2 control?

Release of cytochrome C into the cytosol (regulates the intrinsic pathway).

What are the two types of Bcl2 family proteins?

Anti-apoptotic (pro-survival; blocks cytochrome C release) and pro-apoptotic (pro-death; promotes release).

What are the BH domains of anti-apoptotic proteins?

4 domains: BH4, BH3, BH1, BH2 (e.g., Bcl2, Bcl-XL).

Give an example of a BH123 pro-apoptotic protein.

Bak.

What is a BH3-only protein example?

Bad.

What happens when Bak is activated?

It aggregates in the mitochondrial outer membrane and releases cytochrome C → Apaf1 → apoptosome.

What does MOMP stand for?

Mitochondrial outer membrane permeabilization.

Where do anti-apoptotic Bcl2/BclxL sit and what do they do?

On the cytosolic surface of the outer mitochondrial membrane; they bind pro-apoptotic proteins (BH123; Bak) and prevent aggregation into the active form.