Cytoskeleton, Stem Cells, and Cancer Biology Overview

Cytoskeleton

A complex set of structures that are very dynamic.

The large network consisting of protein fibers and other molecules that gives shape and structure to cells in the body.

what does the cytoskeleton deal with

cell movement

cell shape

organelle movement

organelle orientation

Actin filaments

Also known as microfilaments, they are the smallest type of filament measuring 7-9 nm.

double helix

f-actin

Intermediate filaments

Medium size filaments measuring 10 nm

toughest filament

long, unpolarized, and less dynamic.

serves as a rope in desmosomes and nucleus

Microtubules

The largest filament

measuring 25 nm,

appears during mitosis in the metaphase plate and axons.

Deforming force

The force needed to snap the filaments.

Actin deformation

Actin had low deformation and medium deforming force.

Microtubule deformation

Microtubules had high deformation and low deforming force.

Intermediate filament strength

Intermediate filaments had the best deformation and deforming force, proving that it is the strongest.

Cytochalasin D (toxin to filaments)

A toxin that inhibits polymerization by binding to the + end.

Phalloidin (toxin to filaments)

comes from death cap mushrooms

inhibits depolymerization

there arent antibodies for filaments so we can use phalloidin for staining filaments

treatment for phalloidin consumption

eating raw meat. raw meat attacks the filaments in the meat and not your cells

Thymosin beta 4 (polymerization regulator)

A sequestering protein that regulates ATP G actin binding

works with profilin to regulate polymerization

Profilin (polymerization regulator)

An ATP/ADP exchanger that promotes swapping ADP for ATP G actin.

Cofilin (polymerization regulator)

A severing protein that severs off a chunk of F actin from the - end by adding ADP.

Capping proteins ((polymerization regulator)

Proteins that bind to F actin ends to prevent treadmilling.

Cap Z

A capping protein that binds to the + end of F actin.

Tropomodulin

A capping protein that binds to the - end of F actin.

Formin (polymerization regulator)

dimer

nucleating protein that starts nucleation.

Actin

makes up 10% of all proteins in a cell

alpha actin (actin subtypes)

contractive structures (aka filopodia)

Beta actin (actin subtypes)

leading edge structures

Gamma actin (actin subtype)

stress fibers

G-actin (globular)

Globular actin that requires ATP and Mg2+.

F-actin (filament)

Filamentous actin that is decorated with myosin S1 and appears as an arrowhead on TEM imaging.

pointed end if - side

barbed end is + side

Treadmilling

A process where G actin is added and removed at either ends, resulting in no net increase or decrease in size once steady state is established.

+ end experiences treadmilling 5x faster than - end

polymerization of G actin like this forms f-actin

Critical concentration

A concentration level akin to chemical equilibrium in the context of actin polymerization.

optical traps/tweezers

uses infrared light to determine force of f actin filaments

opsonization and phagocytosis

require f actin to form the shape of the WBC phagocytosing

Listeria

A bacteria that is non-mobile until it reaches the cell

causing severe food poisoning

nucleates f actin b/c it has f actin tail

can cross placenta in pregnant women

becomes motile by binding to f actin alongside act A

types of intermediate filaments

keratin (skin)

desmin

neurofilaments

lamin

-nucleus

iimportant for karyo skeleton

Epidermolysis bullosa simplex

disease caused by defective keratin filaments in basal lamina

causes susceptibility to shear stress and recurrent skin infections

dystrophic epidermolysis bullosa (DEB)

“Butterfly children” with skin as fragile as a butterfly wing

VYJUVEK

A treatment that topically transfects COL7A1 gene to code for Collagen VII to restore skin keratin.

Microtubules structure

Made up of 13 protofilaments

Building blocks of microtubules

alpha beta tubulin

-dimer

-alpha tubulin has non- exchangeable GTP

-beta tubulin has exchangeable GTP

have pigments and that is what lets animals become camoflaged

ex. flounder melanosis

MAPS

Microtubule associated proteins

MAP2

A microtubule associated protein.

Tau

Associated with Parkinson's and Alzheimer's; keeps microtubules intact.

XMAP215 and CLASP

Increases polymerization of microtubules.

Kinesin 13

Increases depolymerization of microtubules.

Katanin

Aids in synaptic pruning.

Colchicine (drug)

Promotes microtubule depolymerization; used to treat cancer and gout.

Taxol/Taxotere (drug)

Enhances polymerization of microtubules; treats cancer by inducing apoptosis.

Kinesin (molecular motor)

Used in anterograde transmission from - to + end.

Dynein (molecular motor)

Used in retrograde transmission from + to - end.

Kartagener syndrome

Defect in dynein.

STEM cell

A cell that can divide and differentiate; some can divide a certain number of times before differentiating fully.

uses for stem cells

tissue engineering

drug discovery

regenerative medicine

food production

recovery after diseases

“clean meat”

wooly mammoth DNA used to make meat

Differentiation

Cell's ability to form into a specified type of cell.

Progenitor cells

Restricted lineage cells

can only differentiate into 1-2 specific types of cells.

Totipotent

STEM cells that can differentiate into ANY specific cell type.

Chimera test

used to see if stem cells from a given sample are totipotent or not

Pluripotent

Can differentiate into MANY cell types.

Multipotent

Differentiates into LESS cell types.

Unipotent

Differentiates into only ONE cell type.

Transdifferentiation “DIRECT REPROGRAMMING”

When a differentiated cell differentiates into another cell type without first reverting to an embryonic step

happens in the prescence of transcription factors

Dedifferentiation

When a differentiated cell turns into an embryo-like cell

can be induced by chemicals like reversine.

Redifferentiation

When embryo-like cells differentiate into a new cell type

this is how lizards and newts can regenerate limbs and eye lenses.

STEM cell niche

STEM cell microenvironment critical to control cell division and differentiation.

what is a stem cell niche composed of

neighboring cells

extracellular matrix

growth factors

environmental factors (pH, oxygen, tension, pressure)

Blastocyst

Early stage fertilized egg where hESCs are derived from.

Homing

STEM cells knowing where to go

ex. damaged tissues release factors for endogenous MSCs to home.

iPSCs (Induced pluripotent STEM cells;)

iPSC reprogramming factors introduced to somatic cells to bring them to embryonic like stage.

Fusogenic

STEM cells can randomly fuse together to form tetraploid STEM cells

can lead to cancer.

stem cells can undergo fusion when injected into human due to the stress of putting them into human

Bioethics

Moral code on bio research that vary country by country.

Therapeutic cloning

Embryonic cloning to use the cells for therapeutic purposes.

-diabetes

-alzheimers

-injury

embryo developed in vitro (lab)

Reproductive cloning

Making genetically identical organisms

in vivo

Adult STEM cells

Found in adipose.

adipose derived mesenchymal stem cells

Fetal STEM cells

Stem cells derived from fetal tissues.

amniotic

umbilical

placental

Embryonic stem cells

Stem cells derived from embryos, also known as hESCs and hPSCs.

Induced pluripotent STEM cells

Stem cells that are reprogrammed from adult cells, not used for research in the US but used in Japan and Australia.

SCID mice

Mice with Severe Combined Immunodeficiency, used to test if a candidate STEM cell can differentiate in vivo.

Somatic cell nuclear transfer (SCNT)

A method of generating STEM cells in vitro

somatic cell nucleus from chronically sick person placed into embryo to differentiate, cells removed and placed back into the person

Pro of somatic nuclear transfer

autologous stem cell therapy

-donor is the receiver

Con of somatic cell nuclear transfer

ethics?

low success rate

Sir Ian wilmut (successful clones)

dolly the sheep

John Gurdon

cloned frogs before wilmut

nobel prize winner

little Nicky (2004) (successful clones)

cloned cat

first successful cloning of primates

2018

Parthenogenesis

A reproductive process where no sperm is needed for pregnancy.

happens in sea urchins and starfish

hPSCs

Cells that arise from parthenogenesis.

pros/cons of hPSCs

pro: if we did this it would only take 200-300 eggs to be able to get hPSCs for anyone on earth

con: all alleles would be homozygous b/c of no sperm=higher chance of expressing mutation

induced pluripotent stem cells (iPSCs)

master genes

adding 4 of them to a human adult cell makes an iPSCs

they reverse the age of genes

discovered by Shinya Kamanaka and James Thomason

(HOX, Oct3/4, Sox2, Nanog)

Tumorigenicity

The potential of STEM cells to form tumors, specifically teratocarcinomas

stem cells have long telomeres

Immunogenicity

potential for activation of immune response

more frequent stem cell injections raises the chance of anaphylaxis (allergic reaction)

Inappropriate differentiation

The risk of STEM cells differentiating into the wrong cell type.

ex. woman got stem cells near her eye for tissue replacement and stem cells differentiated into bone instead

C. Elegans

A multicellular roundworm that serves as a model organism

simple anatomy

short life cycle

well mapped genome

mass reproduction

Benefits of using C. Elegans

Easy to grow on agar, non-pathogenic, and useful for discovering RNAi and apoptotic genes.

Anucleate cells

Cells that do not have a nucleus.

Skin cells

Only basal lamina cells have a nucleus; the rest serve as a barrier so there is no need for nucleus

Lens fibers

Type of fiber cell that is anucleate.

RBCs enucleate

Red blood cells remove their nuclei.

Nuclear envelope

Outer part of the nucleus

made up of 50% nuclear pores

-Allow diffusion for proteins less than 62kDa.

Nucleoplasmin

Pentameres with each subunit being 33kDa; total of 165kDa.

Functions of Nucleoplasmin

First chaperone protein discovered

nucleosome assembly

genome stability

transcription regulation.

Lamin

Intermediate filaments found in the nucleus only

interconnect chromatin to form nuclear skeleton

Types of Lamin

Three main types: A, B, and C.