HETEROCYST - Cyanobacteria - page 218

13) GENES REGULATING THE PATTERN
i) patA (all0521) : During isolation and characterization of Tn5-induced mutants, Liang et al . (1992)
recognized a PAT-1 mutant that differentiated heterocysts at the terminal positions. The cloned
fragment did not contain Tn5 but had a frame-shift mutation due to introduction of a single adenine
in a stretch of eight adenine residues at positions 316-323. Due to this, there was termination in
the reading frame, 40 nucleotides downstream of the shift point. To recognize the particular gene
sequence from Anabaena genome, a cosmid library of genes of Anabaena sp. strain PCC 7120 was
used for complementation. The complementing gene sequence (1.1kb) restored the wild-type
phenotype and was designated as patA (for pattern). patA was then disrupted by the introduction
of a Sp-Sm resistance cassette and then transferred into Anabaena sp. strain PCC 7120. Single and
double recombinants were obtained. Cloning and sequence analysis of patA gene and its deduced
amino acid sequence revealed it to be a protein with 379 amino acids belonging to the group of
response regulators. The C-terminal region of PatA is a phosphoacceptor (receiver) domain bearing
resemblance to CheY of E . coli. patA gene is required for the formation of intercalary heterocysts and
in the patA mutants the pattern of heterocyst formation is thus disturbed because of the formation
of only terminal heterocysts. patA mutation also suppresses Mch-phenotype when hetR gene is
overexpressed (Liang et al ., 1992). Orozco et al . (2006) created a patA null mutant (UHM101) that
differentiates only terminal heterocysts resembling the patA mutant phenotype described previously
(Liang et al ., 1992). A patA hetN double mutant (UHM112, patA deletion mutant with hetN expression
under petE promoter) exhibited delayed Mch-phenotype. Unlike the patA deletion mutant which
differentiated only terminal heterocysts, under inductive conditions in presence of copper UHM112
differentiated multiple heterocysts at terminal positions. In addition, they created a double deletion
mutant ( patA - and patS - ) that had the expression of hetN under the petE promoter. The Mch-phenotype
of this mutant is distinctive for the reason that in addition it differentiated multiple terminal heterocysts
thus decreasing suffi ciently the interheterocyst distance. This suggests that the inactivation of patS is
mainly responsible for such a phenotype. Makarova et al . (2006) reported a highly conserved N-terminal
domain in PatA protein and termed it as PATAN (after PatA N-terminus) and a potential helix-turn-
helix domain, characteristic of proteins that bring about protein-protein interactions associated with
development and differentiation in cyanobacteria. Studies on transcriptional regulation of patA gene
revealed the presence of three tsps at -305, -614 and -645 nucleotide positions. Transcription from tsp
position -305 occurred in all cells of Anabaena sp. strain PCC 7120 after nitrogen step-down whereas
transcription from tsps -614 and -645 positions occurred in proheterocysts. Introduction of P patA - gfp
construct showed GFP fl uorescence at the site of cell division and overexpression of patA resulted in
the formation of enlarged cells. It means that the N-terminal and C-terminal domains of PatA function
independently and patA gene has a role in cell division (Young-Robbins et al ., 2010). The expression of
patA in the heterocysts after 3 to 6 h of nitrogen step-down of Anabaena sp. strain PCC 7120 is probably
because of the presence of two conservative NtcA-binding sites in the promoter region of patA and its
interaction with NtcA (Bastet et al ., 2010).
ii) patB : Another suspected Fix - mutant, isolated and characterized by Liang et al. (1993) by Tn5-
induced mutagenesis of Anabaena sp. strain PCC 7120, PAT-2 exhibited a delay of ~21 h in heterocyst
formation with a decreased interheterocyst distance of 3 to 8 cells in the older cultures resulting
in increased heterocyst frequency. The cloned fragment from PAT-2 mutant did not reveal the
interposed Tn5 but contained a deletion of G at nucleotide 1342 position resulting in a frame-shift
mutation. This caused the deletion of 62 amino acid C-terminal helix-turn-helix portion from the
protein. Studies on complementation of the gene from the cosmid lirary of Anabaena sp. strain