Discuss the following statement: The basal lamina of muscle fibers serves as a molecular

Given the numerous processes inside cells that are regulated by changes in Ca2+ concentration, it seems likely that Ca2+-dependent cellcell adhesions are also regulated by changes in Ca2+ concentration.

Tight junctions perform two distinct functions: they seal the space between cells to restrict paracellular flow and they fence off plasma membrane domains to prevent the mixing of apical and basolateral membrane proteins.

The elasticity of elastin derives from its high content of helices, which act as molecular springs.

Integrins can convert mechanical signals into intracellular molecular signals.

Comment on the following (1922) quote from Warren Lewis, who was one of the pioneers of cell biology. Were the various types of cells to lose their stickiness for one another and for the supporting extracellular matrix, our bodies would at once disintegrate and flow off into the ground in a mixed stream of cells.

Cell adhesion molecules were originally identified using antibodies raised against cell-surface components to block cell aggregation. In the adhesion-blocking assays, the researchers found it necessary to use antibody fragments, each with a single binding site (so-called Fab fragments), rather than intact IgG antibodies, which are Y-shaped molecules with two identical binding sites. The Fab fragments were generated by digesting the IgG antibodies with papain, a protease, to separate the two binding sites (Figure Q191). Why do you suppose it was necessary to use Fab fragments to block cell aggregation?

The food-poisoning bacterium Clostridium perfringens makes a toxin that binds to members of the claudin family of proteins, which are the main constituents of tight junctions. When the C-terminus of the toxin is bound to a claudin, the N-terminus can insert into the adjacent cell membrane, forming holes that kill the cell. The portion of the toxin that binds to the claudins has proven to be a valuable reagent for investigating the properties of tight junctions. MDCK cells are a common choice for studies of tight junctions because they can form an intact epithelial sheet with high transepithelial resistance. MDCK cells express two claudins: claudin-1, which is not bound by the toxin, and claudin-4, which is. When an intact MDCK epithelial sheet is incubated with the C-terminal toxin fragment, claudin-4 disappears, becoming undetectable within 24 hours. In What we dont know What are the regulatory mechanisms that control the rearrangement of cellcell junctions in epithelia during early development? What roles do mechanical force and tension play in these rearrangements? How do extracellular matrix proteins and carbohydrates influence the localization and actions of extracellular signal molecules or their cell-surface receptors? How do intracellular adaptor proteins coordinate the activation of integrin proteins and their interactions with cytoskeletal components and their response to changes in mechanical force acting on cellmatrix junctions? Given that extracellular matrix molecules have the ability to present ordered arrays of signals to cells, might the exact spatial relationships between such signals carry a message beyond that of the individual signals themselves? Figure Q191 Production of Fab fragments from IgG antibodies by digestion with papain (Problem 196). Problems p19.01/19.01 sites of papain cleavage PAPAIN sites for antigen binding IgG antibody Fab fragments 1088 Chapter 19: Cell Junctions and the Extracellular Matrix Figure Q192 Effects of Clostridium toxin on the barrier function of MDCK cells (Problem 197). (A) Addition of toxin from the basolateral side of the epithelial sheet. (B) Addition of toxin from the apical side of the epithelial sheet. For a given voltage, a higher resistance (ohms cm2) gives less paracellular current. the absence of claudin-4, the cells remain healthy and the epithelial sheet appears intact. The mean number of strands in the tight junctions that link the cells also decreases over 24 hours from about four to about two, and they are less highly branched. A functional assay for the integrity of the tight junctions shows that transepithelial resistance decreases dramatically in the presence of the toxin, but the resistance can be restored by washing out the toxin (Figure Q192A). Curiously, the toxin produces these effects only when it is added to the basolateral side of the sheet; it has no effect when added to the apical surface (Figure Q192B). A. How can it be that two tight-junction strands remain, even though all of the claudin-4 has disappeared? B. Why do you suppose the toxin works when it is added to the basolateral side of the epithelial sheet, but not when added to the apical side?

It is not an easy matter to assign particular functions to specific components of the basal lamina, since the overall structure is a complicated composite material with both mechanical and signaling properties. Nidogen, for example, cross-links two central components of the basal lamina by binding to the laminin -1 chain and to type IV collagen. Given such a key role, it was surprising that mice with a homozygous knockout of the gene for nidogen-1 were entirely healthy, with no abnormal phenotype. Similarly, mice homozygous for a knockout of the gene for nidogen-2 also appeared completely normal. By contrast, mice that were homozygous for a defined mutation in the gene for laminin -1, which eliminated just the binding site for nidogen, died at birth with severe defects in lung and kidney formation. The mutant portion of the laminin -1 chain is thought to have no other function than to bind nidogen, and does not affect laminin structure or its ability to assemble into the basal lamina. How would you explain these genetic observations, which are summarized in Table Q191? What would you predict would be the phenotype of a mouse that was homozygous for knockouts of both nidogen genes?

Discuss the following statement: The basal lamina of muscle fibers serves as a molecular bulletin board, in which adjoining cells can post messages that direct the differentiation and function of the underlying cells.

The affinity of integrins for matrix components can be modulated by changes to their cytoplasmic domains: a process known as inside-out signaling. You have identified a key region in the cytoplasmic domains of IIb3 integrin that seems to be required for inside-out signaling (Figure Q193). Substitution of alanine for either D723 in the chain or R995 in the chain leads to a high level of spontaneous activation, under conditions where the wild-type chains are inactive. Your advisor suggests that you convert the aspartate in the chain to an arginine (D723R) and the arginine in the chain to an aspartate (R995D). You compare all three chains (R995, R995A, and R995D) against all three chains (D723, D723A, and D723R). You find that all pairs have a high level of spontaneous activation, except D723 vs R995 (the wild type) and D723R vs R995D, which have low levels. Based on these results, how do you think the IIb3 integrin is held in its inactive state?

The glycosaminoglycan polysaccharide chains that are linked to specific core proteins to form the proteoglycan components of the extracellular space are highly negatively charged. How do you suppose these negatively charged polysaccharide chains help to establish a hydrated gel-like environment around the cell? How would the properties of these molecules differ if the polysaccharide chains were uncharged?

At body temperature, L-aspartate in proteins racemizes to D-aspartate at an appreciable rate. Most proteins in the body have a very low level of D-aspartate, if it can be detected at all. Elastin, however, has a fairly high level of D-aspartate. Moreover, the amount of D-aspartate increases in direct proportion to the age of the person from whom the sample was taken. Why do you suppose that most proteins have little if any D-aspartate, while elastin has levels of D-aspartate that increase steadily with age?

Your boss is coming to dinner! All you have for a salad is some wilted, day-old lettuce. You vaguely recall that there is a trick to rejuvenating wilted lettuce, but you cannot remember what it is. Should you soak the lettuce in salt water, soak it in tap water, or soak it in sugar water, or maybe just shine a bright light on it and hope that photosynthesis will perk it up?

A plant must be able to respond to changes in the water status of its surroundings. It does so by the flow of water molecules through water channels called aquaporins. The hydraulic conductivity of a single aquaporin is 4.4 1022 m3 per second per MPa (megapascal) of pressure. What does this correspond to in terms of water molecules per second at atmospheric pressure? [Atmospheric pressure is 0.1 MPa (1 bar) and the concentration of water is 55.5 M.]