A Novel Endo-(beta)-Mannanase Gene in Tomato LeMAN5 Is Associated with Anther and Pollen Development

Endo-(beta)-mannanase (EC 3.2.1.78) is involved in cell wall disassembly and the weakening of plant tissues by degrading mannan polymers in the cell walls. Endo-(beta)-mannanase genes are expressed in tomato (Lycopersicon esculentum) seeds (LeMAN1 and LeMAN2) and fruits (LeMAN3 and LeMAN4). A novel endo-(beta)-mannanase gene (termed LeMAN5) was found in the tomato genome by genome-walking PCR and bacterial artificial chromosome library screening. The 5ʹ-upstream region of this endo-(beta)-mannanase gene contained four copies of the pollenspecific cis-acting elements POLLEN1LELAT52 (AGAAA). A GUS-reporter gene driven with the putative LeMAN5 promoter (-543 to +38) was activated in anthers and pollen of transgenic Arabidopsis, with the highest (beta)glucuronidase activity detected in pollen. (beta)-Glucuronidase expression was detected in mature pollen retained in sporangia, discharged pollen, and elongating pollen tubes in transgenic Arabidopsis. Consistently, expression of LeMAN5 mRNA and endo-(beta)-mannnanase activity was detected in tomato anthers and pollen. In anthers, the highest mRNA expression and endo-(beta)-mannanase activity were detected during late stages of anther development, when pollen maturation occurred. Endo-(beta)-mannanase activity was present in discharged pollen, which was easily eluted in a buffer, indicating that the enzyme proteins are probably secreted from, and deposited on, the surface of pollen. These data suggest that the LeMAN5 endo-(beta)-mannanase is associated with anther and pollen development.