Chloroplast Division Protein ARC3 Regulates Chloroplast FtsZ-Ring Assembly and Positioning in Arabidopsis through Interaction with FtsZ2

Abstract

Chloroplast division is initiated by assembly of a mid-chloroplast FtsZ (Z) ring comprising two cytoskeletal proteins, FtsZ1 and FtsZ2. The division-site regulators ACCUMULATION AND REPLICATION OF CHLOROPLASTS3 (ARC3), MinD1, and MinE1 restrict division to the mid-plastid, but their roles are poorly understood. Using genetic analyses in Arabidopsis thaliana, we show that ARC3 mediates division-site placement by inhibiting Z-ring assembly, and MinD1 and MinE1 function through ARC3. ftsZ1 null mutants exhibited some mid-plastid FtsZ2 rings and constrictions, whereas neither constrictions nor FtsZ1 rings were observed in mutants lacking FtsZ2, suggesting FtsZ2 is the primary determinant of Z-ring assembly in vivo. arc3 ftsZ1 double mutants exhibited multiple parallel but no mid-plastid FtsZ2 rings, resembling the Z-ring phenotype in arc3 single mutants and showing that ARC3 affects positioning of FtsZ2 rings as well as Z rings. ARC3 overexpression in the wild type and ftsZ1 inhibited Z-ring and FtsZ2-ring assembly, respectively. Consistent with its effects in vivo, ARC3 interacted with FtsZ2 in two-hybrid assays and inhibited FtsZ2 assembly in a heterologous system. Our studies are consistent with a model wherein ARC3 directly inhibits Z-ring assembly in vivo primarily through interaction with FtsZ2 in heteropolymers and suggest that ARC3 activity is spatially regulated by MinD1 and MinE1 to permit Z-ring assembly at the mid-plastid.

The Endosomal Protein CHARGED MULTIVESICULAR BODY PROTEIN1 Regulates the Autophagic Turnover of Plastids in Arabidopsis

“…Impaired division is observed in plants overexpressing FTSZ1 by <3-fold (Stokes et al, 2000;Schmitz et al, 2009). Overexpression of several other plastid division proteins also causes related but distinct abnormalities in chloroplast division, FTSZ filament organization, and chloroplast morphology (Vitha et al, 2003;Maple et al, 2007;Zhang et al, 2009Zhang et al, , 2013. By contrast, overexpression of PDV1 and/or PDV2 has the opposite effect in that it results in an increased plastid division rate and smaller and more numerous plastids per cell (Okazaki et al, 2009).…”

Roles of Arabidopsis PARC6 in Coordination of the Chloroplast Division Complex and Negative Regulation of FtsZ Assembly

“…Recently, S. pombe has emerged as a valuable heterologous system in which to analyze the behavior of bacterial and chloroplast FtsZ filaments as well as to test the interactions with and effects of putative FtsZ assembly regulators (Srinivasan et al, 2007(Srinivasan et al, , 2008TerBush and Osteryoung, 2012;Zhang et al, 2013). When expressed alone in S. pombe, PARC6 77-573 -mCherry fluorescence appeared diffusely localized in the cytosol (Fig.…”

“…Furthermore, genetic analysis suggested that, unlike ARC6, which positively regulates FtsZ assembly (Vitha et al, 2003), PARC6 functions partly as a negative regulator of FtsZ assembly. Interaction assays provided evidence that this negative regulation may be mediated by interaction of the N terminus of PARC6 with the stromal division protein ARC3 (Pyke et al, 1994;Shimada et al, 2004;Maple et al, 2007), a Z-ring positioning factor recently shown to inhibit Z-ring assembly and/or promote FtsZ filament and Z-ring destabilization (TerBush and Osteryoung, 2012;Zhang et al, 2013;Johnson et al, 2015). Although the interaction of PARC6 with FtsZ was not detected previously, the significance of this finding has remained uncertain in the absence of definitive data on PARC6 topology ).…”

In situ structure of FtsZ mini-rings in Arabidopsis chloroplasts

“…Changes in FtsZ expression levels or assembly lead to dramatic phenotypes such Open Access *Correspondence: holzen@tamu.edu 4 Microscopy and Imaging Center, Texas A&M University, Interdisciplinary Life Sciences Building, 2257 TAMU, College Station, TX 77843-2257, USA Full list of author information is available at the end of the article as single giant chloroplasts covering entire mesophyll cells [8][9][10]. Z-ring formation/assembly depends on the balance between FtsZ and accessory chloroplast division proteins, which either stabilize (ARC6 [11,12], MinE [3,13,14]) or destabilize (ARC3 [15], PARC6 [16]) FtsZ assemblies.…”

“…The arrangement of FtsZ protofilaments in the Z-ring in chloroplasts and how they bend and proceed in chloroplast division remains unknown. Highly curved, small ring-like FtsZ assemblies, termed mini-rings, have been reported in plant chloroplasts and studied by optical microscopy [3,15,30,31]. It is hypothesized that the structural signature principles identified in these highly curved small assemblies may also play a role in the actual Z-ring.…”