Chloroplast development requires the coordinated expressions of nuclear and chloroplast genomes, and both anterograde and retrograde signals exist and work together to facilitate this coordination. We have utilized the Arabidopsis yellow variegated (var2) mutant as a tool to dissect the genetic regulatory network of chloroplast development. Here, we report the isolation of a new (to our knowledge) var2 genetic suppressor locus, SUPPRESSOR OF VARIEGATION9 (SVR9). SVR9 encodes a chloroplast-localized prokaryotic type translation initiation factor 3 (IF3). svr9-1 mutant can be fully rescued by the Escherichia coli IF3 infC, suggesting that SVR9 functions as a bona fide IF3 in the chloroplast. Genetic and molecular evidence indicate that SVR9 and its close homolog SVR9-LIKE1 (SVR9L1) are functionally interchangeable and their combined activities are essential for chloroplast development and plant survival. Interestingly, we found that SVR9 and SVR9L1 are also involved in normal leaf development. Abnormalities in leaf anatomy, cotyledon venation patterns, and leaf margin development were identified in svr9-1 and mutants that are homozygous for svr9-1 and heterozygous for svr9l1-1 (svr9-1 svr9l1-1/+). Meanwhile, as indicated by the auxin response reporter DR5:GUS, auxin homeostasis was disturbed in svr9-1, svr9-1 svr9l1-1/+, and plants treated with inhibitors of chloroplast translation. Genetic analysis established that SVR9/SVR9L1-mediated leaf margin development is dependent on CUP-SHAPED COTYLEDON2 activities and is independent of their roles in chloroplast development. Together, our findings provide direct evidence that chloroplast IF3s are essential for chloroplast development and can also regulate leaf development.
The development of functional chloroplasts relies on the fine coordination of expressions of both nuclear and chloroplast genomes. We have been using the Arabidopsis () () leaf variegation mutant as a tool to dissect the regulation of chloroplast development. In this work, we screened for genetic enhancer modifiers termed () mutants and report the characterization of the first locus, We showed that encodes the cytosolic 80S ribosome 40S small subunit protein RPS21B and the loss of causes the enhancement of leaf variegation. We further demonstrated that combined S21 activities from EVR1 and its close homolog, EVR1L1, are essential for Arabidopsis, and they act redundantly in regulating leaf development and leaf variegation. Moreover, using additional cytosolic ribosomal protein mutants, we showed that although mutations in cytosolic ribosomal proteins all enhance leaf variegation to varying degrees, the 40S subunit appears to have a more profound role over the 60S subunit in regulating VAR2-mediated chloroplast development. Comprehensive genetic analyses with suppressors that are defective in chloroplast translation established that the enhancement of leaf variegation by cytosolic ribosomal protein mutants is dependent on chloroplast translation. Based on our data, we propose a model that incorporates the suppression and enhancement of leaf variegation, and hypothesize that VAR2/AtFtsH2 may be intimately involved in the balancing of cytosolic and chloroplast translation programs during chloroplast biogenesis.
Protein homeostasis is essential for cellular functions and longevity, and the loss of proteostasis is one of the hallmarks of senescence. Autophagy is an evolutionarily conserved cellular degradation pathway and is critical for the maintenance of proteostasis. Paradoxically, autophagy deficiency leads to accelerated protein loss by unknown mechanisms. We discover that ABS3 subfamily of multidrug and toxic compound extrusion (MATE) transporters promote senescence in natural and carbon-deprivation conditions in Arabidopsis thaliana . The senescence-promoting ABS3 pathway functions in parallel with the longevity-promoting autophagy to balance plant senescence and survival. Surprisingly, ABS3 subfamily MATE proteins interact with ATG8 at late endosome to promote senescence and protein degradation without the canonical cleavage and lipidation of ATG8. This non-autophagic ATG8-ABS3 interaction paradigm is likely conserved among dicots and monocots. Our findings uncover a previously unknown non-autophagic function of ATG8 and an unrecognized senescence regulatory pathway controlled by the ATG8-ABS3-mediated proteostasis.
Summary Histone H2B monoubiquitination (H2Bub1) is recognized as a crucial eukaryotic regulatory mechanism that controls a range of cellular processes during both development and adaptation to environmental changes. In Arabidopsis, the E2 conjugated enzymes UBIQUITIN CARRIER PROTEINs (UBCs) ‐1 and ‐2 mediate ubiquitination of H2B. Here, we elucidated the functions of UBC1 and ‐2 in salt‐stress responses and revealed their downstream target genes. Real‐time quantitative PCR assays showed that UBC1 and ‐2 positively regulated the salt‐induced expression of MYB42 and Mitogen‐Activated Protein Kinase 4 (MPK4). Chromatin immunoprecipitation assays revealed that H2Bub1 was enriched weakly on the chromatin of MYB42 and MPK4 in the ubc1,2 mutant. We further determined that UBC1/2‐mediated H2Bub1 enhanced the level of histone H3 tri‐methylated on K4 (H3K4me3) in the chromatin of MYB42 and MPK4 under salt‐stress conditions. MPK4 interacted with and phosphorylated MYB42. The MPK4‐mediated MYB42 phosphorylation enhanced the MYB42 protein stability and transcriptional activity under salt‐stress conditions. We further showed that MYB42 directly bound to the SALT OVERLY SENSITIVE 2 (SOS2) promoter and mediated the rapid induction of its expression after a salt treatment. Our results indicate that UBC1 and ‐2 positively regulate salt‐stress responses by modulating MYB42‐mediated SOS2 expression.
Plant interphase cortical microtubules (cMTs) mediate anisotropic cell expansion in response to environmental and developmental cues. In Arabidopsis thaliana, KATANIN 1 (KTN1), the p60 catalytic subunit of the conserved MT‐severing enzyme katanin, is essential for cMT ordering and anisotropic cell expansion. However, the regulation of KTN1‐mediated cMT severing and ordering remains unclear. In this work, we report that the Arabidopsis IQ67 DOMAIN (IQD) family gene ABNORMAL SHOOT 6 (ABS6) encodes a MT‐associated protein. Overexpression of ABS6 leads to elongated cotyledons, directional pavement cell expansion, and highly ordered transverse cMT arrays. Genetic suppressor analysis revealed that ABS6‐mediated cMT ordering is dependent on KTN1 and SHADE AVOIDANCE 4 (SAV4). Live imaging of cMT dynamics showed that both ABS6 and SAV4 function as positive regulators of cMT severing. Furthermore, ABS6 directly interacts with KTN1 and SAV4 and promotes their recruitment to the cMTs. Finally, analysis of loss‐of‐function mutant combinations showed that ABS6, SAV4, and KTN1 work together to ensure the robust ethylene response in the apical hook of dark‐grown seedlings. Together, our findings establish ABS6 and SAV4 as positive regulators of cMT severing and ordering, and highlight the role of cMT dynamics in fine‐tuning differential growth in plants.
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