could you please answer these questions that I attached here about cell functions????????????…..
1. ErbB-2 is a receptor found on the plasma membrane of mammalian cells. Recent studies have
shown that this protein can move from the plasma membrane to the nucleus. Because most
proteins that shuttle between the cytoplasm and the nucleus are soluble and not integral membrane
proteins, the mechanism by which ErbB2 undergoes this transport is of particular interest (Giri et
al., Mol. Cell Biol. 25:11005, 2005)
In the first experiment, RNA interference (siRNA) is used to reduce expression levels of importin 1.
Cells are either not subjected to siRNA (), or have their expression of importin reduced (imp),
or are subjected to a nonsense siRNA (NS). The presence of ErbB-2 and two other proteins, tubulin
and histone H3, in both cytoplasmic and nuclear fractions is then analyzed by immunoblotting using
antibodies against each of these proteins. The data are shown on the immunoblot below in Figure
(a). In the second experiment, lysates of cells expressing either wild-type (WT) ErbB-2 or mutant
ErbB-2 lacking its nuclear localization signal (NLS) are immunoprecipitated with antibody to
ErbB-2 or a control antibody (mIgG). The immunoprecipitates are then examined by
immunoblotting with antibodies to importin 1 or to ErbB-2 as shown in Figure (b).
between ErbB-2 and importin 1?
What do these data suggest about the mechanism of localization of tubulin and histone H3?
In further studies, cells were either untreated () or were induced to overexpress wild-type Ran
(WT) or mutant Ran in which a glutamine at residue 69 is replaced with a leucine (Q69L). Ran
Q69L hydrolyzes GTP poorly. The nuclear and cytoplasmic fractions are assessed by
immunoblotting for the presence of ErbB-2, importin 1, tubulin, and histone H3, as shown at the
left in Figure (c). The amount of Ran and ErbB-2 in the total cell lysates is also shown in Figure (c).
From the data provided, what model can you propose to explain the mechanism by which
ErbB-2 is translocated into the nucleus?
Cells are treated with the drug leptomycin B (LMB) which inhibits the nuclear export protein Crm1,
which is an exportin, and then analyzed for the localization of ErbB2, as shown by
immunofluorescence in Figure (d). Regions of overlap between nuclear and ErbB-2 staining appear
white, some of which are indicated by white arrowheads. In addition, nuclei are isolated at 0, 2, and
12 hours after LMB addition and assessed for the presence of ErbB-2, as shown in Figure (e).
What effect does LMB have on ErbB-2 localization? What would you predict would happen
if LMB were used in cells overexpressing Ran Q69L?
2. Many organisms make a single type of clathrin heavy chain and a single type of clathrin light chain;
thus they make a single kind of clathrin coat. How is it, then, that a single clathrin coat is used for
several different transport pathways, such as Golgi to early endosomes and plasma membrane to
early endosomes, that each involves different specialized cargo proteins?
3. 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. Why do you suppose it was necessary to use Fab fragments to block