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Functional analysis and characterization of a water deficit stress induced gene lp5 in loblolly pine (Pinus taeda L.)
xii, 106 leaves : illustrations ; 28 cm.
Investigations to isolate and characterize genes induced by water deficit stress (WDS) in loblolly pine resulted in several WDS responsive clones obtained by differential screening of a cDNA library (Chang et al 1995). Of these, lp5 was found to encode a putative glycine-rich protein (GRP) based on sequence similarity. In the present study, we were interested in analyzing the expression, regulation and induction of this clone in detail. It has been shown that the 5' flanking sequences of the lp5 gene were able to drive the expression of the uidA gene encoding β-glucuronidase (GUS) in transgenic tobacco. The experiments established that the upstream sequences of lp5 provide for selective expression in vascular tissues and determined a minimal promoter of 210 bp. The data did not show any significant induction due to WDS, possibly due to the fact the experiments were carried out in a heterologous system. The role of induced LP5 was assessed by cloning it behind a 35S promoter to overexpress the protein and assess changes in morphology, cell wall elasticity and tolerance to WDS in transgenic tobacco. Data obtained from pressure-volume curve statistics did not show significant changes in elasticity nor in WDS tolerance. By employing immunocytochemistry, pine tissues were examined for putative localization of the protein encoded by lp5. Antibodies generated against the LP5 were able to localize the protein in the cell walls of vascular tissues in primary xylem and secondary xylem, and the deposition was found to be induced upon imposition of WDS. The results from the present investigation led us to suggest that lp5 is spatially and differentially regulated, and mediated by developmental and stress signaling. This study is a significant step forward in understanding the molecular mechanisms involved in the regulation, expression and functioning of the first grp gene in pine.
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Includes bibliographical references (leaves 90-101).