slide set 15 Flashcards
integrins what are they and what do they bind
membrane protein that binds the basal lamina
integrins structure
dimer of alpha and beta subunits
24 types in human (8 beta, 18 alpha)
integrins bind extracellularly to ECM proteins and intracellularly to linker proteins that attach to actin
important linker protein
talin!
focal adhesions
clusters of multiple integrins bound to talin, linking to bundles of actin filaments (stress fibers)
assembly
assembly and disassembly must occur in the right space at the right time for motility
how do we study the role of focal adhesions in cell migration?
- fluorescently label a protein found in focal adhesions (EX: vinculin) and observe localization over time
- in video, cell moves with polarity
ECM aka
extracellular matrix
cells have to make and release attachments to ECM
- Outside-in signaling
- integrin attachment to ECM triggers actin attachment on the inside
- Inside-out signaling
- signals stimulate actin and associated proteins to associate with integrins
- this changes conformation of the extracellular domain to bind ligand
active vs inactive integrin
inactive (weak binding) state (folded)
active (strong binding state) (unfolded)
differences are due to changes in structure
used data from X-ray crystallography
many integrins cluster together to form….
strong focal adhesions!
many other proteins cluster on the cytosolic side
(some bind to integrin tail, actin, or other proteins)
some of the proteins that make up a focal adhesion are enzymes that can detect tension generated by attachment to the ECM (more tension stabilizes the complex and forms larger adhesions
FAK aka
focal adhesion kinase
when is FAK active
FAK is active when cells are attached
talin and mechanical sensing
talin contributes to mechanical sensing by integrin-based cell-matrix adhesions
tension causes talin to unfold partially and expose multiple sites to bind vinculin
vinculin binds actin filaments, which creates more tension and anchors for actin filaments
talin is a tension sensor
Extracellular matrix (ECM)
cells synthesize and secrete the ECM and then organize it into a larger structure
talin positive feedback loop
tension leads to binding leads to more tension
ECM includes….
- bone, teeth
- tendon
- ligament
- cornea
- cartilage
- basal lamina
- connective tissues
- the proteins that make up the ECM provide strength and support, but also function in cell signaling (chemical and mechanical)
3 broad categories of ECM proteins
proteoglycans and GAGs
fibrous proteins
glycoproteins
basal lamina
thin layer of ECM that separates epithelial cells from the connective tissue
prevents fibroblasts from migrating into epithelial tissue, but immune cells can cross
cells on both sides secrete the proteins that make up the basal lamina
multiple components of ECM proteins are assembled to form it
components vary but most include laminin, type IV collagen, and sometimes fibronectin
glycoproteins
laminin is the primary organizer of the basal lamina
other examples: nidogen, fibronectin
laminin anchors proteins to cells
image is electron micrographs of laminin molecules shadowed with platinum
structural model of basal lamina attached to a cell’s integrins
other type of glycoproteins in ECM
fibronectin, CYR61, tenascin, thrombospondin (don’t memorize)
many are large scaffold protein with multiple copies of specific protein-interaction domains
proteoglycans
hyaluronan, perlecan, decorin, aggrecan
a protein core that is covalently attached to a sugar (specifically a glycosaminoglycan [GAG])
sugars provide hydration to tissues
the sugars are negatively charged, which attracts positively charged water molecules to form a hydrated gel
GAG sugars are not branched (unlike previous sugars we discussed)
fibrous proteins
EX: type IV collagen, fibrillar collagen
collagens are major proteins of ECM
formed of 3 polypeptide chains, each called an alpha chain
alpha chains wrap around each other to form a stiff triple-stranded helix
collagen molecules associate together into….
fibrils and fibers
how do cells organize collagens
cells pull on collagens and organize them
2 pieces of chick embryo heart were cultured on a collagen gel
individual fibroblast cells migrate on the collagen, pulling on in and shaping it into fibers between the 2 pieces in the process
ECM degradation
cells must degrade the ECM
ECM need to be degraded for normal turnover of proteins, remodeling of the ECM and for cells to migrate past it
extracellular proteases degrade ECM
