A Putative O-Linked β-
N
-Acetylglucosamine Transferase Is Essential for Hormogonium Development and Motility in the Filamentous Cyanobacterium Nostoc punctiforme
Authors: Khayatan Behzad , Bains Divleen K. , Cheng Monica H. , Cho Ye Won , Huynh Jessica , Kim Rachelle , Omoruyi Osagie H. , Pantoja Adriana P. , Park Jun Sang , Peng Julia K. , Splitt Samantha D. , Tian Mason Y. , Risser Douglas D. 
Published: February 2017

ABSTRACT

Most species of filamentous cyanobacteria are capable of gliding motility, likely via a conserved type IV pilus-like system that may also secrete a motility-associated polysaccharide. In a subset of these organisms, motility is achieved only after the transient differentiation of hormogonia, which are specialized filaments that enter a nongrowth state dedicated to motility. Despite the fundamental importance of hormogonia to the life cycles of many filamentous cyanobacteria, the molecular regulation of hormogonium development is largely undefined. To systematically identify genes essential for hormogonium development and motility in the model heterocyst-forming filamentous cyanobacterium
Nostoc punctiforme
, a forward genetic screen was employed. The first gene identified using this screen, designated
ogtA
, encodes a putative O-linked β-
N
-acetylglucosamine transferase (OGT). The deletion of
ogtA
abolished motility, while ectopic expression of
ogtA
induced hormogonium development even under hormogonium-repressing conditions. Transcription of
ogtA
is rapidly upregulated (1 h) following hormogonium induction, and an OgtA-GFPuv fusion protein localized to the cytoplasm. In developing hormogonia, accumulation of PilA but not HmpD is dependent on
ogtA
. Reverse transcription-quantitative PCR (RT-qPCR) analysis indicated equivalent levels of
pilA
transcript in the wild-type and Δ
ogtA
mutant strains, while a reporter construct consisting of the intergenic region in the 5′ direction of
pilA
fused to
gfp
produced lower levels of fluorescence in the Δ
ogtA
mutant strain than in the wild type. The production of hormogonium polysaccharide in the Δ
ogtA
mutant strain is reduced compared to that in the wild type but comparable to that in a
pilA
deletion strain. Collectively, these results imply that
O
-GlcNAc protein modification regulates the accumulation of PilA via a posttranscriptional mechanism in developing hormogonia.

IMPORTANCE
Filamentous cyanobacteria are among the most developmentally complex prokaryotes. Species such as
Nostoc punctiforme
develop an array of cell types, including nitrogen-fixing heterocysts, spore-like akinetes, and motile hormogonia, that function in dispersal as well as the establishment of nitrogen-fixing symbioses with plants and fungi. These symbioses are major contributors to global nitrogen fixation. Despite the fundamental importance of hormogonia to the life cycle of filamentous cyanobacteria and the establishment of symbioses, the molecular regulation of hormogonium development is largely undefined. We employed a genetic screen to identify genes essential for hormogonium development and motility in
Nostoc punctiforme
. The first gene identified using this screen encodes a eukaryotic-like O-linked β-
N
-acetylglucosamine transferase that is required for accumulation of PilA in hormogonia.