The de novo production of specific proteins, most notably ADH, in seedlings in response to anaerobic stress is a well documented phenomenon (Sachs and Freeling 1978, MGG 161:111; Ferl et al. 1978, MGG in press). Our protein labeling studies have led us to an examination of RNA production under the same conditions.
Studies monitoring the incorporation of P32 or H3 RNA precursors into specific RNAs (as displayed in a variety of high resolution polyacrylamide gel systems and detected by autoradiography) have produced some very interesting results. The most notable among these concerns an RNA with an apx. mw of .8 x 106 dalton. Basically the facts on this particular RNA are as follows:
--It is an induced RNA species. That is, it does not appear in RNA preps from unstressed seedling roots labeled in air, but does appear in root preps from identical seedlings labeled early during anaerobic stress induced by a N2 atmosphere.
--Pulse label time course experiments show that this induced RNA is made predominantly during the early stages of anaerobic stress, mostly in the first 8-10 hrs. Some slight production of this species has been noted at later times.
--This RNA is fairly stable. Pulse-chase experiments have shown that this RNA, after being labeled for a 2 hr period early during anaerobic stress treatment, maintained its label after 22 further hours of anaerobic chase in cold medium.
--This RNA is apparently poly A containing, as judged by its ability to reversibly bind to oligo (dT) cellulose. In fact, it appears to be by far the major poly-A containing species that takes up labeled precursor during anaerobic stress.
All the facts mentioned above are consistent with the possibility that this RNA is the messenger RNA for ADH. Although all the correlations fit, to date direct evidence to support this contention is lacking. Currently our efforts to test this possibility are two-fold: 1) We have available many Adh1 nulls, the Adh1-FCm duplication, an Adh1 molecular weight variant U725 and other Adh1 and Adh2 mutant types. These are all being tested in the hope that one or more of them may produce either no or a detectably altered form of this particular RNA. 2) In vitro translation is being used in an attempt to identify the protein product of this RNA and its similarity, if any, to ADH.
Studies concerning the effects of anaerobic stress on other RNA species are also in progress.
Rob Ferl and Drew Schwartz
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