Effect of ethylene on mRNA abundance of three β-galactosidase genes in wild type and mutant tomato fruit

Recent studies indicate that β-galactosidases may play an important role in fruit development and ripening. The temporal expression patterns of a recently discovered family of tomato β-galactosidase (TBG) genes suggest that accumulation of some TBG mRNAs may be regulated by ethylene. Because of the dramatic changes in mRNA abundance patterns that TBG4, TBG5 and TBG6 exhibit at the onset of fruit ripening (when ethylene production begins), we analyzed the effects of ethylene exposure on expression of these genes at 35 days after pollination (Mature Green) in wild type and three tomato ripening-impaired mutants: ripening inhibitor (rin), non ripening (nor) andNeverripe (Nr). RNA gel blot analysis showed that 48 h of ethylene treatment increased accumulation of TBG4 mRNA up to 20-fold, except in the nor mutant, suggesting that the NOR gene product is required for ethylene up-regulation of TBG4. Experiments in which fruit were exposed to ethylene for either 2 or 24 h indicated that the observed TBG4 up-regulation is an indirect response to the ethylene treatment, rather than a primary (or direct) response. Ethylene treatment caused the levels of TBG5 mRNA to decrease 63–93% in wild type and mutant fruit at the Mature Green stage, but had no effect after the onset of ripening. Ethylene dramatically decreased the mRNA abundance of TBG6 in wild type and all mutants at the Mature Green stage. It was shown previously that the TBG4-encoded enzyme has β-galactosidase and exo-galactanase activity. In order to elucidate its potential cell wall modifying activity, the TBG5-encoded enzyme was expressed in yeast. β-Galactosidase and exo-galactanase activity of the TBG5-encoded enzyme were confirmed through a quantified release of galactosyl residues from p-nitrophenyl-β-d-galactopyranoside and from tomato fruit cell wall fractions containing β(1→4)-d-galactan, respectively. The TBG6-encoded product was not successfully expressed in yeast. We present a summary of the potential function of the TBGs studied here and their mRNA accumulation in relation to the climacteric production of ethylene in tomato fruit.