Identification and characterization of the Anabaena sp. strain PCC 7120 hupL DNA rearrangement

Abstract

Programmed DNA rearrangements that occur during cellular differentiation are uncommon and have been described in only two prokaryotic organisms. In Anabaena sp. strain PCC 7120 at least three programmed DNA rearrangements occur during heterocyst differentiation. Heterocysts are terminally differentiated cells specialized for nitrogen fixation. These rearrangements take place late during heterocyst differentiation and involve the deletion of DNA elements from within the nifd, fdxn, and hupl genes. In the hupl rearrangement a 10.5-kb DNA element is excised from within the hupl gene by site-specific recombination between two 16-bp direct repeats that flank the element. The predicted HupL polypeptide is homologous to the large subunit of [NiFe] uptake hydrogenases and to HupU which forms part of the HupTLTV operon and is thought to be involved in regulation of uptake hydrogenase. hupl is expressed similarly to the nitrogen fixation genes; hupl message was detected only during the late stages of heterocyst development. The predicted product of a gene located upstream of hupl, shows sequence similarity to the small subunit of uptake hydrogenase and to HupU. An open reading frame, named xisc, was identified near one end of the hupl DNA element and encodes the recombinase required for excision of the hupl element. Inactivation of xisc in the chromosome of Anabaena PCC 7120 prevents rearrangement of the hupl element. Expression of xisc in E. coli is sufficient to cause site-specific rearrangement of an artificial substrate. The predicted XisC protein shows sequence similarity to XisA, the recombinase required for excision of the nifd element, and to the catalytic domain of the Int family of site-specific recombinases. Site-directed mutagenesis of the invariant Arg3O8 and Tyr433 prevented XisC from rearranging the artificial substrate in E. coli.