Field of Science

Interaction of a ∆hfq mutation with the rpoD mutation

Here's the last part of the summary of what our senior co-op technician has been doing.  The last set of experiments tested the interaction between a new ∆hfq mutation we've been studying, which reduces competence) and the rpoD mutation (which increases competence).

The Hfq protein binds small regulatory RNAs, helping them to form base-pairs with the mRNAs that they regulate.  In other bacteria we know that this base pairing can either reduce the mRNA expression (usually mediated by RNase E degradation) or increase it (by reducing the effect of otherwise-inhibitory mRNA secondary structure).  Our ∆hfq mutation reduces competence in MIV-induced cells by about 10-fold, suggesting that it increases the translatability of sxy mRNA.

The technician tested whether this effect is still seen in cells with the rpoD mutation.  She first had to construct the double-mutant strain.  This was relatively easy because the ∆hfq mutation is 'marked' with a SpcR cassette, and the honours student who's studying this mutation had already made chromosomal DNA.  So she used his chromosomal DNA to transform strain RR753 (rpoD mutant) to spectinomycin resistance.

She then did a competence time course, following development of competence in rich medium in four strains.  In the graph below we see that the ∆hfq mutant (green line) develops competence later than the wildtype cells (KW20, blue line) and to a lower final level.  This mutation also reduces the competence of the rpoD mutant (compare red and purple lines), although not as severely.

So we conclude that, whatever Hfq is doing to promote competence, it's still at least partly needed by the rpoD mutant.

The honours student has been analyzing the interactions of the ∆hfq mutant with other factors and other hypercompetence mutations.  I'll do a separate post pulling this together, unless he does it on his blog first.

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