Researchersh ave isolated two E-cadherin mutant isoforms that are hypothesized r

Question

Researchersh ave isolated two E-cadherin mutant isoforms
that are hypothesized ro function differently from the isoform
of the wild-type E-cadherin. An E-cadherin negative mammary
carcinoma cell line was transfected with the mutant
E-cadherin genes A (part a in the figure, triangles) or B (part b)
(triangles)o r the wild-type E-cadhering ene( black circles)a nd
( a ) Anti-E-cadherin
20 40
Time (min)
20 40
Time (min)
80
60
40
80
60
E
540
E20
80
60
80
60
( b )
E
540
E20
844 cHAPTER1 9 | TNTEGRATTNCGE LLStN TO TtSSUES
40
compared to untransfected cells (open circles) in an aggregation
assay. In this assay, cells are first dissociated by trypsin
treatment and then allowed to aggregate in solution over a period
of minutes. Aggregating cells from mutants A and B are
presented in panels a and b respectively. To demonstrate that
the observed adhesion was cadherin mediated, the cells were
pretreated with a nonspecific antibody (left panel) or a functionblocking
anti-E-cadherin monoclonal antibody (right panel)'
a. Vhy do cells transfected with the wild-type Ecadherin
gene have greater aggregation than control, untransfected
cells?
b. From these data, what can be said about the function
of mutants A and B?
c. Sfhy does the addition of the anti-E-cadherin monoclonal
antibodS but not the nonspecific antibody, block
aggregation?
d.
'What
would happen to the aggregation ability of
the cellst ransfectedw ith the wild-type E-c^adherigne nei f the
assay were performed in media low in Ca'-?

Explanation / Answer

Answer:

a. E-cadherins are a special class of type-1 transmembrane protein which plays important roles in cell adhesion and helps adherens junctions to bind cells. The E-cadherin has its binding determinants located at the extracellular domain which facilitate cell-cell adhesion and aggregation.

The control untransfected cells expresses basal level of E-cadherins on the cell membrane compared to the wild type where it is overexpressed. So, the wild type has greater aggregation.

b. The mutant A has a lesser degree of aggregation capacity compared to the wild type while the B mutant's aggregation capacity is almost equivalent to the untransfected control cells. Probably the mutations are located in the extracellular domain of the protein leading to a decreased aggregation capacity.

Loss of E-cadherins have been suggested to be responsible for tumor cell invasion, leading to metastasis. So, the mutant B should have more invasive property (due to lesser aggregation) compared to the A mutant.

c. The anti-E-cadherin antibody binds specifically to the E-cadherins present on the surface of the membranes (extracellular domain), thereby blocking the domains and its aggregation properties.

The non-specific antibody can bind randomly all over the cell surface. So, there is very low probability that the cell surface E-cadherins will get blocked.

d. "Cadherins" stands for "Calcium dependent adhesion" and depends upon calcium for their function.

So, the cells wouldn't undergo the E-cadherin mediated aggregation and perform their function in media lacking calcium.

Related Questions

Navigate

• Browse (All)
• Browse R
• Subjects

Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.