Particle-bound phytochrome: Differential pigment release by surfactants, ribonuclease and phospholipase C

Surfactants and hydrolytic enzymes were used to probe the natureof the constituents) to which phytochrome binds in paniculatefractions from red-irradiated Cucurbita. ¹⁴C]-choline and ³H]-uridinepre-labelled tissue was used to monitor the release of phospholipidsand RNA by these agents. Ribonuclease (RNase) digestion of 20,000xgpellets eliminates both the phytochrome and ribonucleoprotein(RNP) which cosediment at 31S. Little ¹⁴C]-choline occurs inthe 31S fraction and the amount is not changed by RNase digestion.This is further evidence that phytochrome binds directly tothe RNP in the 31S fraction rather than to any membranous materialpresent. The distribution profile of the RNA in a second ( =‘heavy’)phytochrome fraction does not correlate with that of the pigment.This suggests that the phytochrome in this fraction is not boundto RNP. The RNA is of ribosomal origin but much less degradedthan that of the 31S RNP and is resistant to RNase digestion.Phospholipase C releases>80% of the ¹⁴C]-choline from the‘heavy’ fraction without freeing phytochrome. Thisindicates that the pigment does not bind to the polar head groupsof the membrane phospholipids present. Low concentrations ofdeoxycholate dissociate phytochrome from this fraction withoutreleasing substantial quantities of integral membrane proteinsor phospholipids. Some RNP is dislodged by the surfactant butthe phytochrome and RNP are not released as a complex. The datasuggest that the pigment in the ‘heavy’ fractionmay be loosely bound to a protein constituent rather than toRNP or polar phospholipids. ¹This work was done while on sabbatical leave from the WeizmannInstitute of Science, Rehovot, Israel. (Received April 1, 1976; )