The role of profilin in maize pollen growth and development
Abstract
The small actin-binding protein profilin is found in organisms from all eukaryotic kingdoms and is involved in the regulated reorganization of the actin cytoskeleton. Plants are unique because they have large multigene families of profilin isoforms. The aim of the present work is to establish whether the profilin isoforms in maize have equivalent function or whether they have unique biochemical properties that tailor specific isoforms for specific functions. Maize provides a good system to study this question because at least 5 profilin isoforms are expressed in pollen. Analysis by 2-dimensional gels and immunoassays revealed that the levels of particular isoforms changed dramatically during the development of maize pollen. Furthermore, measurements of total actin, F-actin and profilin concentrations revealed that maize pollen has remarkably low F-actin content. The high levels of profilin in pollen are sufficient to account for such a low concentration of F-actin. The approach to determine if these isoforms have different functions was to characterize the biochemical properties of native profilin or recombinant profilin isoforms and to microinject the proteins into plant cells, to compare their effect on an actin dependent process. There are multiple profilin isoforms expressed specifically in pollen and they were all found to have similar affinities for actin and poly- L-proline (PLP) as profilin purified from maize pollen. One profilin isoform that is expressed at low levels in pollen was found to have significantly higher affinity for actin and PLP. When the recombinant profilin isoforms were microinjected into living plant cells the isoform with high affinity for actin was found to be significantly more effective. These results were consistent with the model that actin-binding activity is the primary determinant for causing changes in cell architecture. A mutant of maize profilin 1 that replaced tyrosine at position 6 with phenylalanine was characterized. The mutant profilin had significantly higher affinity for PLP and was unchanged in its affinity for actin compared to the wild-type protein. When this mutant profilin was microinjected into plant cells, it was significantly more effective at altering cell architecture than the wild-type profilin. These results indicate that binding to PLP can modulate profilin function in the complex environment of the cell.
Degree
Ph.D.
Advisors
Staiger, Purdue University.
Subject Area
Cellular biology|Botany|Plant propagation
Off-Campus Purdue Users:
To access this dissertation, please log in to our
proxy server.