Praline metabolism by germinating Lilium longiflorum pollen. I. Labelling of cytoplasmic, wall and culture medium molecules

Abstract

Radioactivity occurs in trithloroacetic acid (TCA)-soluble and precipitable, cytoplasm and salt-washed walls following germination of Lilium longiflorum, cv. 'Ace' pollen in medium containing [¹⁴C]-proline (Pro). Sephadex gel filtration on G-25 through G-100 was employed to determine whether radioactivity in cytoplasm, wall and growth medium from pollen fed [¹⁴C]-Pro or [³H]=Pro plus [¹⁴C]-arafbinose (Ara) was contained within molecules possessing molecular weights of 5,000 to 100,000 daltones or greater. G-25 elution profiles of a crude cytoplasmic fraction (15,000 X g supernatant) from [¹⁴C]-Pro labelled pollen yielded a radioctive void volume peak and a retarded peak. The void volume peak contained hydroxyproline (Hyp), and exhibited a coincidence of [³H]-Pro and [¹⁴C] -Ara labelling when pollen was double labelled with the two isotopes. This peak also contained radioactivity when pollen was germinated in 2-[³H]-myo-inositol. Germination in medium supplemented with 100 µM 2,2'-dipyridyl eliminated radioactivity from 2-[³H]-myo-inositol or [¹⁴C]-,Pro in the peak. Filtratian on G-25 of aTCA-soluble fraction of a salt-extract of walls from [¹⁴C]-Pro labelled pollen resulted in void volume and two retarded peaks. Void volume and two retarded peaks were also obtained upon G-25 filtration of a cellulase-digest of walls from [M]-Pro labeled pollen. The void volume peak contained Hyp, Lys, Gly, Ala, Ser, Glu and Asp acids, Val, Tyr, Leu or lieu and Pro. Sephadex G-90, 75, and 100 elution profiles of cellulasedigests of walls from [³H]-,Pro and [¹⁴C]-Ara labelled pollen yielded radioactive retarded and Hyp-containing void volume peaks with a coincidence of [³H] and [¹⁴C] labelling. Label in the void volume was obtained when either rhozyme P11- or pepsin-digests of walls from [¹⁴C]-Pro labelled pollen were gel filtered on G-50. Paper electrophoresis coupled with paper chromatography of acid hydrolyzates of salt-washed wall fractions demonstrated 15 of the common amino acids. Gel filtration on G-25 of growth medium in which pollen was germinated resulted in two peaks, one of which eluted in the void volume. contained Hyp and excluded during subsequent gel filtration on G-100.