12004:

Overexpression of expansins in plant cells.

Authors:	Link, Bruce, M.(A)Cosgrove, Daniel, J.(A)
Affiliations:	(A): Biology Department, Penn State University
Presenter:	Link, Bruce M., bml111@psu.edu

Expansins are cell wall loosening proteins encoded by a large superfamily of genes, which have been divided into a and b branches. At least some of the members of this superfamily mediate the “acid – growth” response of plants, that is, the induction of wall extension by low pH. We are currently trying to modify expansin gene expression to assess the consequences for cell growth and cell wall properties. Previously we reported on the cloning of six expansin cDNAs from suspension cultures of tobacco BY2 cells. We have shown that BY2 cells exhibit an acid inducible, expansin-mediated growth response, and that they express expansin genes at low levels (Plant Physiol.(1998)118:907-916). We have taken one of the tobacco a-expansin cDNAs (NtExp1) and reintroduced it into lines of BY2 cells under the control of the constitutive CaMV 35S promoter. One hundred and twelve transformed lines were screened for over-expression at the mRNA level. Forty-five of these lines were selected for having mRNA expression levels that were ten to several hundred fold higher than non-transformed controls. When crude cell wall protein from twenty two of these lines was tested for wall extension activity (creep), 20 of the lines showed higher creep activity. Four of these lines were chosen for having twice the creep activity of non-transformed controls. Western blots indicate that the increase in protein levels is more in line with the results of the creep assay than the Northern blots, indicating that there is at least some post transcriptional control of expansins. We have also carried out similar approaches with Arabidopsis, attempting to overexpress AtExp8 under the control of the CaMV 35S promoter. Selected lines of these transgenic plants show a significant expression of the transgene mRNA, but only modest enhancement of expression at the protein level, as assessed by western blotting or wall extension activity assays. These results suggest that translational or post-translational processes limit accumulation of expansin proteins in these experiments.