I have a question about subcellar localization and of proteins. I have
made a vector which expresses a putative viral protein fused to GFP and
transfected this into HeLa cells. Unfused WT GFP is distributed
throughout all cells in a diffuse pattern. My fusion proteins (I have
made both C and N terminal fusions and so far they have similar
phenotypes), on the other hand are mostly distributed in a distinct
punctate pattern. The interesting thing is that in 90% or so of the
cells the fusion protein is exclusively punctate and nuclear, while in
some cells the glowing intracellular dots are pernuclear, and
additionally, in some cells the glowing dots are exclusively
cytoplasmic. In the rare cell there is apparently a nuclear membrane
expression. The dots range in size from tiny to huge (know this is a
very unscientific explanation) with the largest being about 1/6 the
width of a cell and the smallest being at the limits of visibility.
Some cells have hundreds of small dots and some cells have dozens of
small dots and ca. 4-8 very large ones.
My question in simple terms, and pardon my ignorance, is what can this
mean? Are there known cellular proteins which are either exclusively
nuclear or cytoplasmic depending, for example, the cell cycle? Many
protein shuttle between nuleus and cytoplasm, but I don't know of any
which can be in either but in a single cell are in only one compartment
or the other.
I plan to do all the usual colocalization studies to figure out what
these dots represent, but a hint as to what this variation in
localization might mean would be very helpful. (and if the explanation
is an obvious one, I guesss I have been doing too much virology and not
enough cell biology lately!) Thanks.
David N. Levy
University of Alabama at Birmingham
Birmingham, AL 35294-0007
levy at uab.edu
