Carboxymethylcellulase Activity in Lettuce Seeds Prior to Germination

Abstract

Does the endosperm pose a mechanical resistance on embryonic axis (radicle) growth for lettuce seed (achene) germination? To aid answering this question, the cell wall degrading enzyme, carboxymethylcellul a se (CMCase) was extracted and assayed from lettuce seeds imbibed for 0 to 12 h, prior to germination. Measuring the loss of viscosity of carboxymethylcellulose, CMCase activity was high in dry seeds, low after 6 h of imbibition, high after 9 and 10 h, and then reduced again after 12 h. Fractions from Sephadex columns showed CMCase activity in three peaks labeled E1, E2, E3. The greatest change in CMCase activity during imbibition was with E3 (molecular weight of about 40,000 Daltons) and some reduction in E2 (molecular weight about 280,000). The RNA synthesis inhibitor, 6-methyl purine, eliminated CMCase activity when present from 4.5 to 7 h of imbibition and the protein synthesis inhibitor, cycloheximide, eliminated CMCase activity when present between 5.5 and 9 h. Imbibition in darkness lowered CMCase activity while 15 min of light at 3.5 h restored it and 30 min of far-red light at 3 h eliminated it. Increasing the imbibition temperature to 35°C under light reduced activity while under darkness, activity was eliminated under 24 °C and 35°C. CMCase activity was localized in the endosperm surrounding the embryonic axis (micropylar end) of 9 h imbibed seeds. These observations showed that CMCase was active in de grading the cell wall in the endosperm surrounding the radicle, weakening it, prior to radicle protrusion so that the radicle remains undamaged.