Analysis in insect (Chironomus tentans) salivary gland cells of labeled RNA as a function of time after precursor injection and its distance to the nuclear membrane, cytoplasmic zone analysis, could previously be used to demonstrate the presence of short-lasting gradients in newly labeled ribosomal RNA. Since the gradients were sensitive to puromycin, they are likely to be a result of diffusion restriction due to an engagement of the subunits into polysomes. In the present paper the possibility was explored of recording gradients that were caused by labeled subunits in puromycin-resistant associations, which, in all probability, involve the endoplasmic reticulum. It was found that labeled 28 S and 5 S RNA but not 18 S RNA were present in radioactivity gradients lasting for at least 2 days but less than 6 days. The gradients also remained during inhibition of RNA synthesis by actinomycin, and they were completely resistant to puromycin whether given in vivo or in vitro. The semipermanent gradients formed fhere offer a unique parameter for the in vivo study of conditions for formation and maintenance of heavy subunits in puromycin-resistant bonds. An explanation for these and previous results is that the light subunit, although restricted in movement by engagement to polysomes, is nevertheless free to exchange and spread between rounds of translation, whereas at least part of the heavy subunit population is bound to the endoplasmic reticulum and less free to spread. These results offer a good in vivo correlate to previous results on in vitro exchangeability of subunits.

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