Figure 4.

Characterization of 3′ extended non-coding transcripts originating from bidirectional promoters. (A) Northern blots of CUT095 were hybridized with strand-specific probes recognizing top strand transcripts as diagrammed in the schematic figure, detecting transcripts 5′ of CUT095 (P5′, left), CUT095 (P95, middle) or transcripts 3′ of CUT095 (P3′, right). Note the P95 part is the same blot that appears in Figure 1C. (B) Total RNA was hybridized with no oligo (-) or antisense oligos and to the regions of the SUT129 locus indicated in the diagram on top and then treated with RNAse H. The digested samples were then analyzed by northern blotting using a probe against the middle of SUT129 (gray bar). Note that the eSUT levels decrease only with the middle (M) or 3′ oligos. Quantification of the eSUT/SUT ratio of two repeats is given below, error bars denote standard errors. Note that for the M lane, the SUT signal is the combination of both smaller bands. (C) Effect of CF IA inactivation on short and extended non-coding transcripts. RNA from the indicated strains grown at permissive (25°C) or nonpermissive (37°C) temperatures were analyzed by northern blot. SUT and eSUT transcript species are indicated. The membranes were stripped and re-probed to detect the 5S rRNA (5S) as loading control (bottom). Quantification of non-coding transcript species abundance relative to 5S is given in numbers below the images, “B“ signifies that no signal above background was observed.