Exam 3 Flashcards
extracellular matrix (ECM)
fills the space between cells and binds cells and tissues together
tissues are mosty ___
ECM and cells
different ECMs have ___
different types/proportions of fibrous proteins, polysaccharides, adhesion proteins
basal laminae ECM
supports endothelial cells from beneath them;
surrounds muscle cells, fat, and peripheral nerves;
thin/sheet-like;
secreted by endothelial cells or epithelial cells
connective tissue ECM
(bone, tendon, cartilage, adipose, blood) is mostly ECM;
secreted by fibroblasts;
differ in proportions of proteins and polysaccharides
connective tissue:
adipose ECM
less fibrous, mostly ground substance
connective tissue:
tendons ECM
mostly fibrous, less ground substance
connective tissue:
cartilage ECM
contains lots of polysaccharides for cushion
connective tissue:
bone ECM
is hardened by calcium phosphate crystals
fibrous, secreted proteins
include collagen and elastin
fibrous, secreted proteins:
collagen
rope-like triple helix structure;
made of Gly-X-Y repeats;
forms two structures in ECM:
fibril-forming collagen (more rigid) and network-forming collagen (more flexible, less Gly-X-Y repeats)
fibrous, secreted proteins:
elastin
cross linked structure to form network of elastic fibers that stretch and return to original shape;
expand AND contract;
important in arteries and lungs (which need to expand and contract)
polysaccharides
glycosaminoglycans (GAGs) and proteoglycans
polysaccharides:
glycosaminoglycans (GAGs)
repeated units of disaccharides;
NAG sugar + acidic sugar;
negatively charged, bind + ions to trap water and form hydrated gels
polysaccharides:
proteoglycans
core protein with up to 100 GAGs attached at serine residues
adhesion proteins
link ECM components together and to cell surfaces;
include fibronectin and laminin
adhesion proteins:
fibronectin
organize stuff in connective tissues with binding sites to link collagen, proteoglycans, and cells
adhesion proteins:
laminin
organize stuff in basal laminae with binding sites to link nidogen (binds collagen), proteoglycans, and cells
interactions between cells and the ECM
integrins;
modifying the ECM (MMPs);
growth factors;
cancer
interactions between cells and the ECM:
integrins
heterodimeric transmembrane proteins that anchor to/connect ECM and cytoskeleton
interactions between cells and the ECM:
modifying the ECM (MMPs)
matrix metalloproteases (MMPs) digest collagens, laminin, and adhesion proteins;
include heparanase and hyaluronidases that degrade GAGs;
heparanase expression usually limited to platelets/immune cells
interactions between cells and the ECM:
growth factors
proteins that regulate cell proliferation, migration, and survival/apoptosis;
bind to cell surface receptors and heparan sulfate proteoglycan
interactions between cells and the ECM:
cancer
cancer cells show heparanase expression;
penetration of basement membrane = invasion and metastasis;
liberation of bound growth factors
cell adhesion molecules (CAMs)
help cells recognize each other and their environment;
cell-matrix adhesion molecules;
cell-cell adhesion molecules, many are transmembrane proteins, stable and transient interactions, many are Ca2+ and Mg2+ dependent;
homophilic and heterophilic binding
types of cell adhesion molecules (CAMs)
cell-cell adhesion is ___
selective;
dissociated cells re-aggregate with cells of the same origin
cell-cell adhesion dissociation by ___
trypsin (proteolytic enzyme) and EDTA (removes Ca2+ and Mg2+) destroys protein-protein interactions that hold cells together
stable cell-cell adhesion
cells are not just passively clumped together;
tight junctions;
adherens junctions and desmosomes;
gap junctions
stable cell-cell adhesion:
tight junctions
maintain polarity and barrier function in endo and epithelial cells;
claudins and occludins;
barrier function;
apicobasal polarity;
not very adhesive
stable cell-cell adhesion:
tight junctions claudins and occludins
homophilic binding;
link to cytoskeleton inside cell
stable cell-cell adhesion:
tight junctions barrier function
between plasma membranes of adjacent cells;
blood brain barrier;
zo connects claudin/occludin to actin
stable cell-cell adhesion:
tight junctions apicobasal polarity
active transport of glucose by intestinal epithelial cells from apical domain to basolateral domain
stable cell-cell adhesion:
tight junctions not very adhesive
so need help from adherens and desmosomes
stable cell-cell adhesion:
adherens junctions and desmosomes
indirectly link the cytoskeletons of adjacent cells;
cadherins, classical and desmosomal
stable cell-cell adhesion:
adherens junctions and desmosomes - cadherins
Ca2+ binding increases rod like structure in extracellular domain;
homophilic binding;
classical and desmosomal;
decrease in Ca2+/cadherin mutations cause dead embryos (8-16 cells)
stable cell-cell adhesion:
adherens junctions and desmosoms - classical cadherins
anchored intracellularly by catenins to actin filaments
stable cell-cell adhesion:
adherens junctions and desmosomes - desmosomal cadherins
anchored intracellularly to intermediate filaments
stable cell-cell adhesion:
gap junctions
provide direct cytoplasmic connections between adjacent cells;
connexins and connexon
stable cell-cell adhesion:
gap junctions connexin and connexon
6 connexins form hexamer with channel called connexon that aligns with another cells connexon to form gap junction;
pore for ions and small molecules to diffuse (cAMP, Ca2+, ATP)
transient cell-cell adhesion
cells with nomadic lifestyle and interact transiently (like WBCs);
selectins (inflammation);
integrins (Ig superfamily CAMs)
transient cell-cell adhesion:
selectins
bind cell surface carbohydrates;
L-selectin: WBCs;
P-selectin: platelets;
E-selectin: inflammation activated endothelial cells
transient cell-cell adhesion:
inflammation
endothelial cells express E-selectin then bind to carbohydrates on WBCs and platelets;
initially weak interactions rolling along endothelium until integrins come to help
transient cell-cell adhesion:
integrins
expressed by blood cells;
bind strongly to endothelial CAMs to allow tissue entry between endothelial cells;
immunoglobulin (Ig) superfamily CAMs
transient cell-cell adhesion:
integrins Ig superfamily CAMs
biggest gene family in human genome;
extracellular Ig domain;
transmembrane and GPI-anchored;
homophilic or heterophilic binding;
abundant in immune and neural tissues (ICAM, NCAM)
cell-cell adhesion modification
neural tube formation;
cell migration during development;
synapse formation during development and remodeling
cell signaling scheme
ligand from one cell (transport and type);
received by target cell (location of receptor);
intracellular events (fast vs slow changes);
change in cell function (reversible vs irreversible)
methods of cell signaling
juxtacrine (direct);
endocrine;
paracrine;
autocrine
juxtacrine
direct contact between cells, does not depend on diffusion of the ligand;
gap junctions (metabolic or electrical coupled);
two forms of ligands: secreted ECM molecules or ligand on surface
juxtacrine:
secreted ECM molecules (ligand)
integrins (receptor);
migrating cells travel via ECM molecules (like roads)
juxtacrine:
ligand on surface (ligand)
CAM or cell surface receptor (receptor);
notch signaling in many developmental processes, disrupted in many cancers
endocrine
hormones secreted by endocrine cells travel to target cells via circulatory system;
target cells must express the receptor;
two types of ligands: peptide hormones and lipid-soluble steroid hormones
endocrine:
peptide hormones (ligand)
bind to cell surface receptors (receptor);
include insulin, FSH, prolactin;
synthesized as longer propeptides in ER, cleaved to active form in golgi, packaged into secretory vesicles and released
endocrine:
lipid-soluble steroid hormones (ligand)
bind to intracellular (nuclear) receptors (receptor);
“nuclear” because most are transcription factors that change activity when signal arrives;
derived from cholesterol from ER;
include testosterone, estrogen, progesterone, corticosteroids, thyroid hormone, vitamin D3, retinoids
paracrine
molecules secreted by cells into immediate environment;
diffusion and degradation;
target cells must express the receptor;
synapses (neurotransmitters and neuropeptides);
growth factors (steroid hormones and peptides)
paracrine:
synapse - neurotransmitters (ligand)
typically ligand-gated ion channels (receptor);
they are hydrophilic;
include acetylcholine, GABA, glutamate, dopamine, etc
paracrine:
synapse - neuropeptides (ligand)
cell surface receptors (receptor);
synthesized as polypeptides in ER, cleaved to active form in golgi, and secreted;
include NPY, endorphins, somatostatin, etc
paracrine:
growth factors (ligand)
cell surface receptors (receptor);
regulate proliferation, differentiation, and survival BUT without being directly energetic;
include steroid hormones and peptides;
some work through paracrine, BUT also can be endocrine or autocrine
paracrine:
growth factors (ligand) - peptides
most are synthesized in the ER, post-translationally modified, and shipped out by the golgi in secretory vesicles;
some cytokines;
include hedgehog family, WNT family, etc
autocrine
molecules secreted by cell into immediate environment BUT affects itself;
feedback loop;
cancer
autocrine:
feedback loop
to regulate the secreting cell’s signal
autocrine:
cancer
cancer cells can use autocrine signaling to respond to their own cues
signal transduction
transmits the chemical or physical signal from the surface to the cell interior
G protein coupled receptors (GPCRs)
largest family of cell surface receptors, found in all euks;
transmembrane proteins with 7 membrane spanning α helices;
ligand binds at extracellular N-terminus or loops;
coupled to intracellular signaling pathways
GPCRs:
ligand binds at extracellular N-terminus or loops
small molecules;
peptides;
fatty acids;
light
GPCRs:
coupled to intracellular signaling pathways
cAMP pathway;
phosphatidylinositol pathway;
employ second messengers which trigger intracellular events (ligands are the first messengers)
GPCRs are G protein coupled
rely on G protein cycle;
GPCR is bound to heterotrimeric G protein (α, β, γ);
1) ligand binds GPCR
2) GPCR cytosolic domain changes conformation - gets GEF activity (GDP to GTP, active)
3) α subunit and β/γ dissociate from each other and GPCR (interact with their target enzymes and ion channels)
4) RGSs (regulators of G protein signaling) stimulate GTPase activity of α subunit - gets GAP activity (GTP to GDP, inactive, association with GPCR and β/γ subunits)
hormonal activation of adenylyl cyclase to create cAMP
galpha-s stimulates adenylate cyclase to increase cAMP;
galpha-i inhibits adenylate cyclase to decrease cAMP
secondary messengers
levels affect the activity of downstream proteins;
protein kinase A (PKA);
cAMP regulates ion channels
secondary messengers:
protein kinase A (PKA)
cAMP binds regulatory subunits of cAMP-dependent PKA (protein kinase A), catalytic subunits dissociate and phosphorylate targets
cAMP
increases change activity of cytosolic pathways and nuclear pathways;
impacts skeletal muscle;
catalytic subunits of PKA liberate glucose and FAs and transcription of target genes
cAMP:
catalytic subunits of PKA liberate glucose and FAs
1) catalytic subunits of PKA
2) phosphorylate phosphorylase kinase
3) phosphorylate glycogen phosphorylase
4) breaks down glycogen to glucose-1-P
cAMP:
catalytic subunits of PKA transcription of target genes
1) catalytic subunits of PKA to nucleus
2) phosphorylates CREB (CRE-binding protein)
3) recruitment of co-activators to CREB bound at CREs (cAMP response elements)
4) transcription of target genes
*long term changes
regulation of signal transduction
RSGs (regulators of G protein signaling);
cAMP phosphodiesterases;
phosphatases
regulation of signal transduction:
cAMP phosphodiesterases
regulate secondary messenger levels
regulation of signal trasnduction:
phosphatases
regulate levels of phosphorylation
tyrosine kinases (TKs)
couple cell surface receptors to intracellular phosphorylation events;
tyrosine phosphorylation used for growth, cell survival, differentiation, metabolism;
ligand binds receptor, kinase activity, auto cross phosphorylation (at tyrosine residues)
levels of phosphorylation downstream of TK signaling
tightly controlled;
protein tyrosine phosphatases remove tyrosine phosphorylation
receptor TKs (RTKs)
transmembrane proteins with a single spanning α helix;
N-terminal;
C-terminal;
peptide growth factors;
activation
receptor TKs (RTKs):
N-terminal
extracellular ligand-binding domain
receptor TKs (RTKs):
C-terminal
cytosolic tyrosine kinase activity domain
receptor TKs (RTKs):
peptide growth factors
send signals to many RTKs
receptor TKs (RTKs):
activation for most
1) ligand binding
2) dimerization of receptor directly or by conformation changes
3) auto (cross) phosphorylation of receptor
4) increased TK activity (phosphorylation)
receptor TKs (RTKs):
activation for most - increased TK activity
increased TK activity (phosphorylation) on cytosolic side and provides binding sites for additional proteins that bind to receptor with SH2 domains
non-receptor TKs (nRTKs)
not transmembrane proteins but receive signals from cell-surface transmembrane receptors;
diverse structures;
activation;
cytoplasmic;
JAK/STAT signaling
non-receptor TKs (nRTKs):
diverse structures
have diverse structures but ALL have catalytic domain with tyrosine kinase activity
non-receptor TKs (nRTKs):
activation for most
indirect;
1) ligand binds non-catalytic receptor
2) activation of nRTK kinase activity
3) phosphorylation of receptor and/or autophosphorylation of nRTK
4) increased nRTK activity (phosphorylation) on cytosolic side and provides binding sites for additional proteins that bind to receptor with SH2 domains
non-receptor TKs (nRTKs):
cytoplasmic
they are “cytoplasmic” in general but also anchored at membrane and nuclear envelope;
covalent lipid modifications anchor to plasma membrane;
non-covalently binds to receptors, other proteins, lipids;
SH2 domains;
integrin-binding domains (FAK family), integrins
non-receptor TKs (nRTKs):
JAK/STAT signaling
downstream of cytokine receptors that receive cytokine and some peptide hormones (also downstream of GPCRs and RTKs with diff effects);
1) ligand binds cytokine receptor
2) receptor dimerization activates JAK (nRTK)
3) autophosphorylation of JAK
4) phosphorylation of cytokine receptors
5) SH2 of STAT proteins bind phosphorylation cytokine receptors
6) JAK phosphorylates STATs
7) STATs dimerize at SH2 domains
8) STAT dimer translocates to nucleus and affects gene expression as a transcription factor
mitogen-activated protein kinase (MAPK)
signaling is a cascade of three protein kinases activated downstream of TKs (ERK, JNK, p38);
usually coupled to RTKs;
MAPK family members are serine/threonine kinases;
1) G protein activation by GEF
2) MAP3K activation by G protein
3) MKK activation by MAP3K
4) MAPK activation by MKK
5) target phosphorylation by MAPK
MAPK signaling downstream of an activated TK:
GEF docking
1-3) RTK is activated by ligand binding and phosphorylated
4) GEFs dock at activated receptors with SH2 domains
G protein activation
GEF docking at the activated receptor brings GEFs in proximity to membrane associated, small, monomeric G-proteins;
GEFs exchange GDP to GTP to active Rho or Ras;
GAPs activate GTPase activity to G protein hydrolyzes GTP to GDP to inactive Rho or Ras
G protein activation:
MAPK signaling, G protein families
MAPK signaling is triggered by the activation of G proteins;
Ras family (ERK);
Rho family (JNK, p38)
MAP3K (MKK kinase) activation
are activated by GTP bound G proteins;
1) GTP-Ras anchored at membrane recruits inactive Raf (MAP3K) from the cytoplasm
2) Raf links through Ras to PM
3) phosphorylation of Raf to active form with kinase activity
MKK (MAPK kinase) activation
MAP3Ks phosphorylate MAPK kinases (MKKs) at serine/threonine residues;
1) inactive MEK (MKK) interacts with KSR (kinase suppressor of Ras) in the cytoplasm
2) activated Raf joins KSR/MEK and phosphorylates MEK
MKK (MAPK kinase) activation:
KSR
KSR is a scaffolding protein that organizes components of the cascade;
ligand binding triggers recruitment of KSR/MEK to the plasma membrane
MKK (MAPK kinase) activation:
MEK dual specificity
MEKs are dual specificity kinases - can work on serine/threonine residues AND tyrosine
MAPK activation
MKK phosphorylation activates MAPK to phosphorylate at nuclear and cytoplasmic targets;
1) activated MEK (MKK) phosphorylates ERK (MAPK)
2) ERKs (MAPK) phosphorylate targets at serine/threonine residues
MAPK activation:
ERK (MAPK) phosphorylation
cascade is specific - ERKs are the only physiological substrate of MEKs;
phosphorylated ERKs dissociate from KSR complex
MAPK activation:
MAPK targets
activated ERKs anchor to cytoskeleton for transport to correct location;
some activated ERKs enter mitochondria (manage cell survival) and some enter nucleus (chromatin changes, nuclear import/export (NUPs of NPC), transcriptional activation and suppression)
MAPK activation:
MAPK targets - nucleus transcriptional activation and suppression example
Elk-1 phosphorylation activates IEG (immediate early gene) expression;
IEGs (TFs) trigger downstream gene expression changes in SRGs (secondary response genes)
