Class I [beta]-1,3-Glucanases in the Endosperm of Tobacco during Germination

Abstract

Rupture of the seed coat and rupture of the endosperm are separate events in the germination of Nicotiana tabacum 1. cv Havana 425 seeds. Treatment with 1 O-5 M abscisic acid (ABA) did not appreciably affect seed-coat rupture but greatly delayed subsequent endosperm rupture by more than 100 h and resulted in the formation of a novel structure consisting of the enlarging radicle with a sheath of greatly elongated endosperm tissue. Therefore, ABA appears to act primarily by delaying endosperm rupture and radicle emergence. Measurements of P-1,3-glucanase activity, antigen content, and mRNA accumulation together with reporter gene experiments showed that induction of class I P-1,3-glucanase genes begins just prior to the onset of endosperm rupture but after the completion of seed-coat rupture. This induction was localized exclusively in the micropylar region of the endosperm, where the radicle will penetrate. ABA treatment markedly inhibited the rate of /3-1,3-glucanase accumulation but did not delay the onset of induction. lndependent of the ABA concentration used, onset of endosperm rupture was correlated with the same P-1,3-glucanase content/seed. These results suggest that ABA-sensitive class I p-1,3-glucanases promote radicle penetration of the endosperm, which is a key limiting step in tobacco seed germination.

Class I β-1,3-Glucanase and Chitinase Are Expressed in the Micropylar Endosperm of Tomato Seeds Prior to Radicle Emergence

“…In agreement with previous results with tobacco (Vö geli-Lange et al, 1994;Leubner-Metzger et al, 1995, ␤-1,3-glucanase mRNA, protein, and enzyme activity were expressed specifically in the micropylar endosperm at an early stage of tomato seed germination. However, in contrast to the case with tobacco seeds, where little chitinase activity was detected in imbibed seeds (Leubner-Metzger et al, 1995), chitinase mRNA, protein, and enzyme activity accumulated in the micropylar region of imbibed and germinating tomato seeds essentially in parallel with ␤-1,3-glucanase.…”

“…However, in contrast to the case with tobacco seeds, where little chitinase activity was detected in imbibed seeds (Leubner-Metzger et al, 1995), chitinase mRNA, protein, and enzyme activity accumulated in the micropylar region of imbibed and germinating tomato seeds essentially in parallel with ␤-1,3-glucanase. Although the majority of chitinase mRNA was present in the endosperm cap tissue, a smaller accumulation of mRNA occurred also in the radicle tips.…”

“…1, A and B). Thus, as in the case of germinating tobacco seeds (Leubner-Metzger et al, 1995), ␤-1,3-glucanase was expressed in the micropylar tissues of tomato seeds just prior to radicle protrusion, but in contrast to the situation in tobacco, coordinate expression of chitinase also occurs in tomato. When tomato class I (GluB) and class II (GluA) ␤-1,3-glucanase cDNAs isolated from tomato leaves (van Kan et al, 1992) were used to screen a germinating tomato seed cDNA library, only plaques hybridizing with the GluB cDNA were identified.…”

“…Although ␤-1,3-glucanase expression is often accompanied by expression of chitinase [1, 4-(N-acetyl-␤-dglucosaminide) glycanohydrolase, EC 3.2.1.14] in pathogenesis-related (PR) responses (see below), only very low chitinase activity was detected in germinating tobacco seeds. The absence of significant chitinase accumulation and the close relationship between ␤-1,3-glucanase expression and completion of germination under diverse conditions led to the hypothesis that ␤-1,3-glucanase contributed to the hydrolysis of cell wall components resulting in endosperm weakening at the site of radicle protrusion (Vö geli-Lange et al, 1994;Leubner-Metzger et al, 1995;LeubnerMetzger and Meins, 1999).…”

“…A prediction of the hypothesis of Leubner-Metzger et al (1995) is that if ␤-1,3-glucanase contributes to tomato endosperm weakening, the force required to puncture the endosperm cap should be reduced coincident with increasing ␤-1,3-glucanase activity. Therefore, we compared the change in puncture force of germinating tomato endosperm caps in water with those imbibed in 100 m ABA.…”