The 32000-dalton (Da) shield protein regulating electron transport in Spirodelu oligorrlrizu is an integral chloroplast membrane polypeptide. It is rapidly synthesized, constituting a major chloroplast-translation product in vivo. Following in vitro translation of spirodela chloroplast RNA in a wheat germ system, a 33 500-Da polypeptide is produced. Synthesis of a 33500-Da protein, associated with the chloroplast membrane. is also seen in vivo, within 2 min of pulse-labeling spirodela with radioactive amino acids. Comparative analyses among these polypeptides reveal: (a) all three are deficient in lysine residues; (b) the two 33 500-Da species have indistinguishable partial proteolytic digestion patterns while that for the 32000-Da protein differs only slightly from them; (c) radioactivity from the 33 500-Da polypeptide is rapidly chased in vivo into the 32 000-Da protein, even in the presence of protein synthesis inhibitors. These results show the 33500-Da proteins synthesized in vitro and in vivo to be the precursor form of the 32 000-Da shield protein in spirodela, with processing commencing only alter completion of the precursor polypeptide chain and insertion into the membrane.

The nucleotide sequence of the NS gene of the human influenza virus A/PR/8/34 was determined and found to be the same length (890 nucleotides) as the NS gene of another human influenza virus A/Udorn/72 and of the avian isolate A/FPV/Rostock/34. Comparison of the sequences of the NS genes of the two human influenza viruses shows an 8.9% difference whereas the NS gene of the avian isolate differs by only 8% from that of the human strain A/PR/8/34. The extensive sequence similarity among these three genes does not support the notion of species specific homology groups among NS genes of avian and human influenza virus strains. The primary sequence of the A/PR/8/34 NS gene is consistent with the findings that the influenza virus NS gene may code for two overlapping polypeptides. In addition, an open reading frame potentially coding for a polypeptide 167 amino acids in length was found in the negative strand RNA of the A/PR/8/34 virus NS gene.

The rapidly turning over, photoinduced thylakoid protein, P-32000, is the main pulse-labeled membrane polypeptide in the chloroplasts of Spirodela oligorrhiza, yet little is known of its physiological function. Two hypotheses are tested: that P-32000 synthesis is necessary for thylakoid biogenesis; that it directly participates in photosynthesis. Spirodela cultures were dissected into expanding and fully mature tissue. Fronds from both developmental stages transcribed a 0.5 x 10' dalton RNA likely to be the message for P-32000. As to the protein itself, synthesis occurred in both types of tissue but was considerably enhanced in the fully mature state. Thus, a purely transient, developmental function for P-32000 during thylakoid biogenesis appears ruled out. Low concentrations of D-threo-chloramphenicol severely suppressed P-32000 synthesis but not its turnover. As a result, fronds depleted in P-32000 were obtained. However, photoassimilation of CO2 remained at 86% of normal in tissue > 80% depleted of P-32000. Thus, P-32000 did not appear to be rate-limiting, suggesting that it does not serve as a direct, integral part of the photosynthetic pathway.other procedures which remove more loosely bound proteins (11). P-32000 is itself relatively short lived, exhibiting an order of magnitude of more rapid decay than that of other major plastid polypeptides. Under many conditions there is a close relation between the level of P-33500 template RNA and P-32000 (11, 28). Under certain stress conditions this coordination can be uncoupled (I 1).Plastid membrane proteins of about 32,000 d with properties overlapping those reported in Spirodela have been noted in other genera (5,10,13,15, 23,35). Several studies have also described chloroplast mRNA directed translation of P-32000-like polypeptides in cell-free protein-synthesizing systems (3,20,22). A few communications record the presence of rapidly labeled 0.5 x 106 d chloroplast RNAs not associated with rRNA metabolism products in other plants (18,19,36,38).It is of interest to understand the biological function of the 32,000 d polypeptide. Why is the plastid investing so much of its resources in synthesizing a short lived mRNA and its ephemeral protein product? In this report we test two hypotheses on the function of P-32000: the first, that it is involved in thylakoid biogenesis; and the second, that it forms an integral part of the photosynthetic pathway within the thylakoids.A 32,000 d4 polypeptide, P-32000, is the major membrane protein synthesized within the chloroplasts of the duckweed Spirodela (I 1). An RNA, likely to be the message for P-32000, has a mol wt 0.5 x 106 d (29) turns over rapidly (33), is undermethylated (25) and lacks poly(A) (31,34

ABSTRACTa specific organelle protein; the relationship between chloroplast and surrounding cytoplasm in this control; and the modulation of synthetic control through phylogenetic evolution.The zooflagellate, Euglena gracilis, possesses several attributes which make it an attractive organism for studying chloroplast-related phenomena. In particular, the wild type organism shows a completely reversible transformation of chloroplast to proplastid, with a concomitant shutdown, or induction, of plastid-specific protein synthesis (31,35). In addition, stable mutants which lack chloroplasts and detectable amounts of chloroplast DNA (6) may be readily isolated. As part of our investigation into the synthetic control of specific chloroplast proteins, we present our findings on the purification and char-

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