PRE-MESSENGER-RNAS AND THEIR SPLICING IN THE PRIMARY MEMBRANES OF PEAETIOCHLOROPLASTS - DETECTION BY ELECTRON-MICROSCOPY

Electron-microscopic examination of the primary membranes (PM) isolate d from pea seedling (Pisum sativum L.) etiochloroplasts early in their biogenesis (after a 3-h exposure to light) showed that the principal inclusions within these membranes were thin threadlike networks and ri ngs of various sizes. The abundance of these inclusions coincided in t ime with the peak of transcriptional activity during the PM biogenesis . Many net structures and large rings were formed in parallel-disposed strands. Some circular threadlike structures had a morphology resembl ing that of the maturing transcripts when they underwent splicing, i.e ., the excision of a portion of sequence with its subsequent cyclizati on. This resulted in an accumulation of a great quantity of smaller ri ngs in the PM. The fields of monomeric ribosomes were seen near the la rge rings undergoing splicing. The size and configuration of all the t hreadlike structures corresponded to the morphology of the polyribosom es that would later appear in the PM. The threadlike structures were a ble to be cleared from the electron-transparent part of the membrane b y a trypsin treatment with a subsequent centrifugation through a disco ntinuous sucrose density,gradient. The isolated structures were suscep tible to RNAse treatments. The radioactive label (H-3-UTP) that had be en incorporated into the polynucleotides of the PM was detected in the fractions containing the threadlike structures. We have concluded tha t these structures, already present in the PM before polyribosomes app ear, are the primary transcripts (pre-mRNAs). A new stages of early bi ogenesis of the chloroplast PM has been suggested: (1) a functioning o f the transcriptional complex bound to the PM that results in the appe arance of primary transcripts in these membranes; (2) a processing of the primary transcripts (splicing) into mRNA and rRNA; (3) the assembl y of monomeric ribosomes; and (4) the formation of polyribosomes, whic h are the basis for the further development of the membrane system des cribed in our previous publications.