β-glucuronidases in plants.

In the last decade, the interest for the presence and role of endogenous GUS in plants increased, being previously limited to the use of GUS from E. coli as a reporter gene. Three different GUS genes have been identified in A. thaliana and different roles have been suggested, mainly associated to plant cell wall remodelling and to the regulation of the presence in the active form of molecules with regulative functions. This thesis investigated several aspects: A) artefacts in histochemical GUS detection, B) role of GUS in pollen tube germination and growth, C) GUS expression in N. tabacum and A. thaliana, D) identification of GUS genes in N. tabcum and their phylogenetic analysis in angiosperms. A) Histochemical detection of E.coli GUS activity in transformed plants can be impaired by the presence of GUS inhibitors and by the solubility of an intermediate reaction product formed when X-glu is used as substrate. The expression of LAT52 has been revised. B) The use of saccharolattone, a GUS inhibitor, suggests that GUS is involved in pollen tube germination and growth. C) GUS expression was observed in all organs of N. tabacum and A. thaliana. Expression studies, in collaboration with researchers of Calabria University, have been performed in Arabidopsis, by in situ mRNA hybridization: GUS3 is specifically expressed in border like cells and probably it is involved in their detachment from root tip; GUS1 and GUS2 are expressed in the root cup meristem, resulting perhaps involved in regulation of the mitotic cycle. D) In N. tabacum GUS2 was completely sequenced and one GUS1 and two GUS3 have been partially sequenced. By bioinformatics analysis, GUS genes have been identified in other angiosperms species and a phylogenetic analysis have been conducted.