Pedicel Breakstrength and Cellulase Gene Expression during Tomato Flower Abscission

Abstract

Six cellulase genes were isolated from total RNA of the ethylenetreated tomato (Lycopersicon esculentum Mill.) flower abscission zone by reverse-transcription polymerase chain reaction using degenerate primers to conserved amino acid sequences from known plant cellulases. Four of the gene fragments are homologous to fruit pericarp cellulases. l h e other two are nove1 cellulase genes, referred to as Ce15 and Ce16. Breakstrength and cellulase gene expression were then analyzed in naturally abscising flowers and flower explants. I n both naturally abscising flowers and flower explants induced to abscise in air or ethylene, both new cellulase mRNAs were correlated with flower shedding. Whereas the Ce15 mRNA increased in later stages of abscission, the Ce16 mRNA was present i n nonabscising flowers and then decreased i n the final stage of abscission. A third cellulase, Cell, increased during the final stage of abscission in flower explants and yet did not increase during shedding i n planta, although it was detectable at low levels in all abscission stages. Cell and Ce15 mRNA decreased 99% when indole-3-acetic acid was added during ethylene treatment, consistent with low levels of abscission (3%). I n contrast, Ce16 mRNA increased slightly when indole-3-acetic acid was added. These results suggest that abscission is a multistep process involving both activated and repressed cellulase genes and that the relative importance of each cellulase in the process depends on the physiological conditions under which abscission takes place.Abscission is a process that allows plants to shed an organ from the parent body in a controlled fashion. Separation results from catabolic alterations in the middle lamellae and the primary cell wall in a few layers of specialized cells usually located at the base of the organ to be shed (Sexton and Roberts, 1982). The breakdown of cell-wall components in these specialized cells is so complete that after separation the cells of the scar surface look rounded, as if they were protoplasts (Sexton et al., 1989), and yet the destruction is gradual and spatially confined. These specialized cells have a unique ability to sense and respond to abscission signal(s) by initiating a gene expression program that causes separation (Sexton and Roberts, 1982). The enzymatic machinery engaged by the abscission program operates through severa1 classes of enzymes including endo-P-(1,4)-glucanases, commonly referred to as cellulases (Lewis and Varner, 1970; de1 Campillo et al., 1990),

Immunodetection and Characterization of Tomato Endo-β-1,4-Glucanase Cel1 Protein in Flower Abscission Zones

“…We carried out analysis of changes in BKS in cv VFNT cherry tomato, which produces an abundant supply of flower clusters. In this cultivar flower abscission takes place in a shorter period of time and the average BKS of the pedicel as a whole is smaller than in large tomato cultivars (del Campillo and Bennett, 1996).…”

“…BKS measurements (del Campillo and Bennett, 1996), a quantitative method to monitor the progress of abscission, was also used to examine the relationship between Cell protein accumulation and the progression of abscission. BKS is the force necessary to separate the flower at the abscission site and can be measured using a force gauge.…”

“…Although both Cell and Ce12 mRNA are present in abscission zones and in ripening fruit, Cell mRNA is more abundant than Ce12 mRNA in abscission zones, whereas Ce12 mRNA is most abundant in ripening fruit. Lashbrook et al (1994) demonstrated that both RNAs increase in the abscission zone of tomato flower explants concomitantly with abscission; however, the induction of Cell mRNA was not observed during abscission in planta (de1 Campillo and Bennett, 1996). Two additional members of the EGase family, Ce15 and Ce16, have also been identified in abscission zones, with Ce15 being regulated in the latest stages of abscission (de1 Campillo and Bennett, 1996).…”

Two tomato endoglucanases have a function during syncytium development

“…These enzymes belong to glycosyl hydrolase family 9 (Henrissat 1991) and their substrates probably include xyloglucan, integral and peripheral regions of non-crystalline cellulose and possibly glucomannan (Brummell and Harpster 2001). EGases have been shown to be expressed during fruit ripening, senescence, in elongating and expanding cells, which suggests a role in plant development (Lashbrook et al 1994;del Campillo and Bennett 1996;Catala et al 1997;Catala et al 2000).…”

Arabidopsis endo‐1,4‐β‐glucanases are involved in the formation of root syncytia induced by Heterodera schachtii

“…The general role of endo-1,4-bglucanases is known, but the specific function of most of the members is still unclear. The general function is usually linked to processes such as cell wall biosynthesis and modifications, cell elongation and differentiation, organ abscission (Del Campillo and Bennett, 1996) and fruit ripening (Rose and Bennett, 1999). The family consists of two groups: enzymes with a putative signal peptide (a-and b-subfamily) coding for secreted proteins and enzymes with a membrane-anchoring domain (c-subfamily) coding for non-secreted proteins (Del Campillo, 1999).…”