Chloroplast Division and Morphology Are Differentially Affected by Overexpression of FtsZ1 and FtsZ2 Genes in Arabidopsis,

Abstract

In higher plants, two nuclear gene families, FtsZ1 and FtsZ2, encode homologs of the bacterial protein FtsZ, a key component of the prokaryotic cell division machinery. We previously demonstrated that members of both gene families are essential for plastid division, but are functionally distinct. To further explore differences between FtsZ1 and FtsZ2 proteins we investigated the phenotypes of transgenic plants overexpressing AtFtsZ1-1 or AtFtsZ2-1, Arabidopsis members of the FtsZ1 and FtsZ2 families, respectively. Increasing the level of AtFtsZ1-1 protein as little as 3-fold inhibited chloroplast division. Plants with the most severe plastid division defects had 13-to 26-fold increases in AtFtsZ1-1 levels over wild type, and some of these also exhibited a novel chloroplast morphology. Quantitative immunoblotting revealed a correlation between the degree of plastid division inhibition and the extent to which the AtFtsZ1-1 protein level was elevated. In contrast, expression of an AtFtsZ2-1 sense transgene had no obvious effect on plastid division or morphology, though AtFtsZ2-1 protein levels were elevated only slightly over wild-type levels. This may indicate that AtFtsZ2-1 accumulation is more tightly regulated than that of AtFtsZ1-1. Plants expressing the AtFtsZ2-1 transgene did accumulate a form of the protein smaller than those detected in wild-type plants. AtFtsZ2-1 levels were unaffected by increased or decreased accumulation of AtFtsZ1-1 and vice versa, suggesting that the levels of these two plastid division proteins are regulated independently. Taken together, our results provide additional evidence for the functional divergence of the FtsZ1 and FtsZ2 plant gene families.The first identified proteins of the chloroplast division machinery were homologs of the essential bacterial cell division protein FtsZ (Osteryoung and Vierling, 1995; Osteryoung et al., 1998;Strepp et al., 1998). In contrast with most bacterial genomes that contain only a single gene encoding FtsZ, the nuclear genome of Arabidopsis contains at least three FtsZ genes encoding members of two distinct protein families, FtsZ1 and FtsZ2. AtFtsZ1-1 and AtFtsZ2-1, members of the FtsZ1 and FtsZ2 families, respectively, have been shown to be essential for chloroplast division. When the level of either gene is diminished, chloroplast division is inhibited, yielding cells with as few as one very large chloroplast. One important difference between AtFtsZ1-1 and AtFtsZ2-1 is their predicted localization. AtFtsZ1-1 is targeted to the chloroplast, as is a closely related FtsZ protein from pea (Gaikwad et al., 2000), and is thought to be a component of a division ring that forms on the stromal side of the inner envelope membrane. AtFtsZ2-1, in contrast, lacks a chloroplast transit peptide, is not targeted to the chloroplast in vitro, and is hypothesized to be a constituent of a division ring that assembles on the cytoplasmic surface of the outer envelope membrane. The two rings together are postulated to coordinate chloroplast division (Osteryoung et ...

Rice Cytokinin GATA Transcription Factor1 Regulates Chloroplast Development and Plant Architecture

“…There are multiple FtsH genes that have been shown to demonstrate functional redundancy (Zaltsman et al, 2005). While Arabidopsis contains at least two FtsZ homologs (AtFtsZ1-1 and AtFtsZ2-1; Stokes et al, 2000), rice appears to contain only a single FtsZ copy of this ancestral bacterial division gene. Increased production of FtsZ genes results in a significantly reduced number of enlarged chloroplasts, indicating a severe inhibition of chloroplast division (Stokes et al, 2000;Yoder et al, 2007;Schmitz et al, 2009).…”

“…While Arabidopsis contains at least two FtsZ homologs (AtFtsZ1-1 and AtFtsZ2-1; Stokes et al, 2000), rice appears to contain only a single FtsZ copy of this ancestral bacterial division gene. Increased production of FtsZ genes results in a significantly reduced number of enlarged chloroplasts, indicating a severe inhibition of chloroplast division (Stokes et al, 2000;Yoder et al, 2007;Schmitz et al, 2009). Transgenic Arabidopsis plants overexpressing FtsZ1 showed increased chloroplasts in mesophyll cells, but these plants were chlorotic and died as seedlings (Stokes and Osteryoung, 2003).…”

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

“…In Arabidopsis, overexpression of FTSZ1 or FTSZ2 in the wild-type background results in dose-dependent plastid division defects, with plants having fewer, larger chloroplasts than in wild-type, heterogeneous chloroplast morphology, randomly organized FSTZ filaments, and at least in the case of FTSZ1, long interconnecting bridges McAndrew et al, 2001;Raynaud et al, 2004). 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.…”

“…Slides were incubated in blocking buffer (2% nonfat dry milk and 0.05% Tween 20 in PBS, pH 7.4) for 1 h at room temperature, followed by overnight incubation with anti-FTSZ2-1 antibody (1:500; Stokes et al, 2000) in buffer (2% normal goat serum in blocking buffer). After four washes of 10 min in PBST buffer (PBS containing 0.05% Tween), slides were incubated for 3 h with Alexa Fluor 488 goat anti-rabbit IgG (1:500; Invitrogen).…”

Two Mechanosensitive Channel Homologs Influence Division Ring Placement in Arabidopsis Chloroplasts

“…Two nonredundant families of FtsZ genes are found in seed plants (Osteryoung and Vierling, 1995;Osteryoung et al, 1998;Stokes and Osteryoung, 2003;Schmitz et al, 2009), and both FtsZ1 and FtsZ2 localize to a ring-like structure at the division site Vitha et al, 2001;Fujiwara et al, 2008). Plant and bacterial lines under-or overexpressing FtsZ exhibit enlarged chloroplasts and filamentous cells, respectively, though plants with altered levels of FtsZ are otherwise phenotypically normal (Dai and Lutkenhaus, 1992;Stokes et al, 2000;El-Kafafi et al, 2005Liu et al, 2007;Yoder et al, 2007;Schmitz et al, 2009;Karamoko et al, 2011). Arabidopsis thaliana FtsZ proteins have GTPase activity and are capable of forming filaments in vitro (El-Kafafi et al, 2005;Olson et al, 2010;Smith et al, 2010).…”