The Requirement of WHIRLY1 for Embryogenesis Is Dependent on Genetic Background in Maize

Plastid gene expression is essential to embryogenesis in higher plants, but the underlying mechanism is obscure. Through molecular characterization of an embryo defective 16 (emb16) locus, here we report that the requirement of plastid translation for embryogenesis is dependent on the genetic background in maize (Zea mays). The emb16 mutation arrests embryogenesis at transition stage and allows the endosperm to develop largely normally. Molecular cloning reveals that Emb16 encodes WHIRLY1 (WHY1), a DNA/RNA binding protein that is required for genome stability and ribosome formation in plastids. Interestingly, the previous why1 mutant alleles (why1-1 and why1-2) do not affect embryogenesis, only conditions albino seedlings. The emb16 allele of why1 mutation is in the W22 genetic background. Crosses between emb16 and why1-1 heterozygotes resulted in both defective embryos and albino seedlings in the F1 progeny. Introgression of the emb16 allele from W22 into A188, B73, Mo17, Oh51a and the why1-1 genetic backgrounds yielded both defective embryos and albino seedlings. Similar results were obtained with two other emb mutants (emb12 and emb14) that are impaired in plastid protein translation process. These results indicate that the requirement of plastid translation for embryogenesis is dependent on genetic backgrounds, implying a mechanism of embryo lethality suppression in maize.     

Marker rescue from the Nicotiana tabacum plastid genome using a plastid/Escherichia coli shuttle vector

We recently reported an 868-bp plastid DNA minicircle, NICE1, that formed during transformation in a transplastomic Nicotiana tabacum line. Shuttle plasmids containing NICEI sequences were maintained extrachromosomally in plastids and shown to undergo recombination with NICE1 sequences on the plastid genome. To prove the general utility of the shuttle plasmids, we tested whether plastid genes outside the NICE1 region could be rescued in Escherichia coli. The NICE1-based rescue plasmid, pNICER1, carries NICE1 sequences for maintenance in plastids, the CoIE1 ori for maintenance in E. coli and a spectinomcyin resistance gene (aadA) for selection in both systems. In addition, pNICERl carries a defective kanamycin resistance gene, kan*, to target the rescue of a functional kanamycin resistance gene, kan, from the recipient plastid genome. pNICERl was introduced into plastids where recombination could occur between the homologous kan/kan* sequences, and subsequently rescued in E. coli to recover the products of recombination. Based on the expression of kanamycin resistance in E. coli and the analysis of three restriction fragment polymorphisms, recombinant kan genes were recovered at a high frequency. Efficient rescue of kan from the plastid genome in E. coli indicates that NICE 1-based plasmids are suitable for rescuing mutations from any part of the plastid genome, expanding the repertoire of genetic tools available for plastid biology.     

The role of plastid competition in the control of plastid inheritance in the zonal Pelargonium

The inheritance pattern of mutant white plastids was studied in W × W crosses, in which one mutant was highly stable (W^s) and the other unstable (W^u) owing to the spontaneous restitution (mutation) of white plastids to a new green form. Thirty-six selfs and crosses were made within and between three nuclear type I cultivars, transmitting the unstable plastids, and three nuclear type II cultivars, transmitting the stable plastids. The allelic frequencies of the restituted plastids among the progeny were subjected to an analysis of variance which showed that within each nuclear type the three cultivars were rather similar except for some heterogeneity after W^s × W^u plastid crosses. The relative average transmission of the two mutant plastids in these W × W crosses was estimated and compared with their individual transmission in reciprocal crosses in which one parent contained green plastids. In the latter crosses, the green plastids were superior to the mutant plastids and the unstable plastid mutant was only slightly more successful than the stable mutant. But when the mutant plastids competed against each other, the unstable mutant became greatly superior to the stable mutant and comparable to a green normal plastid. A model to explain these results is discussed.     

Chloroplast‐targeted Hsp90 plays essential roles in plastid development and embryogenesis in Arabidopsis possibly linking with VIPP1

The Arabidopsis genome contains seven members of Hsp90. Mutations in plastid AtHsp90.5 were reported to cause defects in chloroplast development and embryogenesis. However, the exact function of plastid AtHsp90.5 has not yet been defined. In this study, albino seedlings were found among AtHsp90.5 transformed Arabidopsis, which were revealed to be AtHsp90.5 co‐suppressed plants. The accumulation of photosynthetic super‐complexes in the albinos was decreased, and expression of genes involved in photosynthesis was significantly down‐regulated. AtHsp90.5 T‐DNA insertion mutants were embryo‐lethal with embryo arrested at the heart stage. Further investigation showed AtHsp90.5 expression was up‐regulated in the siliques at 4 days post anthesis (DPA). Confocal microscopy proved AtHsp90.5 was located in the chloroplasts. Plastid development in the AtHsp90.5 mutants and co‐suppressed plants was seriously impaired, and few thylakoid membranes were observed, indicating the involvement of AtHsp90.5 in chloroplast biogenesis. AtHsp90.5 was found to interact with vesicle‐inducing protein in plastids 1 (VIPP1) by bimolecular fluorescence complementation system. The ratio between VIPP1 oligomers and monomers in AtHsp90.5 co‐suppressed plants drastically shifted toward the oligomeric state. Our study confirmed that AtHsp90.5 is vital for chloroplast biogenesis and embryogenesis. Further evidence also suggested that AtHsp90.5 may help in the disassembly of VIPP1 for thylakoid membrane formation and/or maintenance.