RNA degradosomes exist in vivo in Escherichia coli as multicomponent complexes associated with the cytoplasmic membrane via the N-terminal region of ribonuclease E.

RNase E isolated from Escherichia coli is contained in a multicomponent "degradosome" complex with other proteins implicated in RNA decay. Earlier work has shown that the C-terminal region of RNase E is a scaffold for the binding of degradosome components and has identified specific RNase E segments necessary for its interaction with polynucleotide phosphorylase (PNPase), RhlB RNA helicase, and enolase. Here, we report electron microscopy studies that use immunogold labeling and freeze-fracture methods to show that degradosomes exist in vivo in E. coli as multicomponent structures that associate with the cytoplasmic membrane via the N-terminal region of RNase E. Whereas PNPase and enolase are present in E. coli in large excess relative to RNase E and therefore are detected in cells largely as molecules unlinked to the RNase E scaffold, immunogold labeling and biochemical analyses show that helicase is present in approximately equimolar amounts to RNase E at all cell growth stages. Our findings, which establish the existence and cellular location of RNase E-based degradosomes in vivo in E. coli, also suggest that RNA processing and decay may occur at specific sites within cells.

Population heterogeneity of higher-plant mitochondria in structure and function.

Mitochondria of rapidly developing mungbean seedlings were fractionated into four populations: two density classes, each from a 1500S and a 150S pellet. Each of the four populations exhibited cytochrome c oxidase (COX) activity and contained mitochondrial DNA and cardiolipin; plastid and glyoxysome content were found to be relatively low. Five mitochondrial membrane proteins, COXII/III, ATPase alpha/beta and porin, and a matrix enzyme, manganese superoxide dismutase (MnSOD), were detected by immunoblots in all four populations. Another matrix enzyme, pyruvate dehydrogenase was detected only in the two respiratory-competent 1500S populations. The two 150S populations contained a previously unidentified organelle that lacked demonstrable respiratory capability. This organelle, which we have tentatively referred to as "slow-sedimenting (ss-) mitochondrion", was small in size (below light-optics resolution, 70-300nm, majority < or =200nm) and possessed a peculiar looking boundary membrane, ribosomes, and an occasional prominent electron-dense spot. Characteristically, ss-mitochondria were almost always in contact with a filament-aligned membrane-like structure of varying length. Cristae structure, while undetected in small ss-mitochondria, appeared in larger individuals. Typical mitochondria were found in the denser 1500S population, while the lighter 1500S population consisted of 300-800 nm mitochondria exhibiting a varying degree of size-dependent inner membrane folding. Using electron microscopy (EM) immunolocalization and serial sectioning, we have identified in situ organelles resembling in size and in fine structure the ss-mitochondria, which also exhibit a size-dependent folding of the inner membrane. These results suggest that small ss-mitochondria may undergo a progressive development in situ. Taken together, our findings demonstrate the existence of a pattern of structure-function-coordinated gross heterogeneity among mitochondria. This pattern of mitochondrial heterogeneity, characterized both in isolated mitochondria and in situ, implies that small ss-mitochondria may represent a type of "nascent mitochondria" derived from a yet unidentified mitochondria-propagation mode operating during rapid seedling growth. Mitochondrial division by binary fission, characterized by the appearance of dumbbell-shaped intermediates, was also detected.