ABSTRACT

Gibberellins (GAs) are plant hormones that regulate a wide range of developmental and growing processes in plants. Levels of active GAs (GA1 and GA4) are regulated by biosynthetic enzymes such as GA-20oxidases (GA20ox) and GA 3-oxidases (GA3ox) and catabolic enzymes such as GA 2-oxidases (GA2ox). The role of some GA metabolism genes on GA homeostasis and tobacco (Nicotiana tabacum var. xanthi) development has been studied using reverse genetics approach. Transgenic plants overexpressing a GA3ox gen (35S:PsGA3ox1) showed small phenotypical variations and slight increases in GA1 content, probably because this is a non-limiting enzyme. Additionally, some GA2ox, NtGA2ox3 and -5, increased their expression in these plants and could be responsible for the lack of GA1 accumulation by its conversion to GA8 (inactive form of GA1). Furthermore, the pattern expression of these GA2ox genes in response to GA levels variations, suggests that they could play an important role in GA homeostasis when a small increases in endogenous GA content occurs. Parallel to this, the simultaneous expression of a GA3ox and a GA20ox (35S:PsGA3ox1 x 35S:CcGA20ox1), caused small variations in the phenotype and active GA levels compared to its GA20ox parental. These results suggest a non limiting character of GA3ox on GA metabolism in tobacco.

On the other hand, the role of GA 2-oxidases genes (catabolism genes) was studied using post-transcriptional RNA interference silencing (RNAi). Tobacco GA2ox enzymes are encoded by at least five genes with high redundancy in expression. To obtain multiple silenced transgenic plants, a hairpin construction with a high-conserved sequence for the five GA2ox genes was performed to silence all those genes. Most of the transgenic silenced GA2ox/RNAi plants showed similar phenotypes to those caused by GA over-production (increases in length of hypocotyls, internodes, leaves, fruit peduncles and in final plant height). Additionally, other phenotypic characteristics like inhibition of root length, flowering delay and low fertility were detected in these plants and could be associated to a GA excess. These results suggest the important role of GA2ox genes in the control of GA levels that regulate many processes of tobacco development.

Finally, the role of GAs on flowering induction has been studied in tobacco plants. When the plants approximate flowering, a progressive reduction of GA1 and GA4 levels was observed by its quantification at different developmental stages in the apical shoots. This result excludes these two GAs as flowering-promoting factors in tobacco. Nevertheless, flowering time in tobacco is delayed either by GAs deficiency or excess. Thus, flowering occurs only when GAs levels are within an optimum range, over a certain minimum or beyond a maximum.