Protein gradients on the nucleoid position the carbon-fixing organelles of cyanobacteria

Abstract

Carboxysomes are protein-based bacterial organelles encapsulating key enzymes of the Calvin-Benson-Bassham cycle. Previous work has implicated a ParA-like protein (hereafter McdA) as important for spatially organizing carboxysomes along the longitudinal axis of the model cyanobacterium Synechococcus elongatus PCC 7942. Yet, how self-organization of McdA emerges and contributes to carboxysome positioning is unknown. Here, we identify a small protein, termed McdB that localizes to carboxysomes and drives emergent oscillatory patterning of McdA on the nucleoid. Our results demonstrate that McdB directly stimulates McdA ATPase activity and its release from DNA, driving carboxysome-dependent depletion of McdA locally on the nucleoid and promoting directed motion of carboxysomes towards increased concentrations of McdA. We propose that McdA and McdB are a previously unknown class of self-organizing proteins that utilize a Brownian-ratchet mechanism to position carboxysomes in cyanobacteria, rather than a cytoskeletal system. These results have broader implications for understanding spatial organization of protein mega-complexes and organelles in bacteria.

The McdAB Carboxysome Positioning System is Widespread Among β-cyanobacteria

“…In bacteria, the ParA/MinD family of ATPases have been primarily studied for their ability to segregate genetic cargos, such as chromosomes and plasmids (Reviews in Baxter and Funnell, 2014; Badrinarayanan et al, 2015). Less studied are ParA/MinD family members implicated in the positioning of diverse protein complexes, including those involved in secretion (Viollier et al, 2002; Perez-Cheeks et al, 2012) , chemotaxis (Thompson et al, 2006; Ringgaard et al, 2011; Alvarado et al, 2017) , conjugation (Atmakuri et al, 2007) , cell division (Raskin et al, 1999; MacCready et al, 2017) and cell motility (Youderian et al, 2003; Kusumoto et al, 2008) , as well as protein-based bacterial microcompartments (BMCs), such as the carboxysome (Savage et al, 2010; MacCready et al, 2018) . For partitioning plasmids, the ParABS system is the best characterized to date.…”

“…In our recent study, we identified a new self-organizing ParA-type ATPase system, McdAB, which is responsible for equidistantly positioning the carbon-fixing organelles of cyanobacteria, carboxysomes (MacCready et al, 2018) . Carboxysomes are essential for photoautotrophic growth of cyanobacteria.…”

“…Through these mechanisms, carboxysomes contribute to greater than 25% of global carbon-fixation through atmospheric CO 2 assimilation (Rae et al, 2013) . Using the model rod-shaped cyanobacterium Synechococcus elongatus PCC 7942, we identified a small novel protein, McdB, responsible for emergent oscillatory patterning of ATP-bound McdA on the nucleoid (MacCready et al, 2018) . While McdB had no identifiable sequence similarities with any known ParB-family members, we found that McdB localized to carboxysomes through multiple shell protein interactions and removed McdA from the nucleoid in their vicinity (MacCready et al, 2018) ; observations that are analogous to ParA patterning following ParB binding to the parS site of plasmids.…”

“…Using the model rod-shaped cyanobacterium Synechococcus elongatus PCC 7942, we identified a small novel protein, McdB, responsible for emergent oscillatory patterning of ATP-bound McdA on the nucleoid (MacCready et al, 2018) . While McdB had no identifiable sequence similarities with any known ParB-family members, we found that McdB localized to carboxysomes through multiple shell protein interactions and removed McdA from the nucleoid in their vicinity (MacCready et al, 2018) ; observations that are analogous to ParA patterning following ParB binding to the parS site of plasmids. Thus, like plasmids, we showed that carboxysomes utilize a Brownian-ratchet based mechanism whereby McdB-bound carboxysome motion occurs in a directed and persistent manner towards increased concentrations of McdA on the nucleoid (Vecchiarelli et al, 2014; Hu et al, 2017; MacCready et al, 2018) (Figure 1C) .…”

“…Interestingly however, our neighborhood analysis around the carboxysome operon in the distantly-related cyanobacterium Gloeobacter kilaueensis JS1 identified a ParA-type ATPase that possessed the signature lysine residue absent in S. elongatus McdA and a small downstream coding sequence. The protein product of this gene loaded onto S. elongatus carboxysomes, which was surprising because it had no sequence homology to S. elongatus McdB, (Figure 8AB: MacCready et al, 2018) . The findings suggest a more rigorous gene neighborhood analysis was necessary to identify other McdAB systems.…”