Non-homologous chromosome pairing during meiosis in haploid Brassica rapa.

KEY MESSAGE: Cytological observations of chromosome pairing showed that evolutionarily genome duplication might reshape non-homologous pairing during meiosis in haploid B. rapa. A vast number of flowering plants have evolutionarily undergone whole genome duplication (WGD) event. Typically, Brassica rapa is currently considered as an evolutionary mesohexaploid, which has more complicated genomic constitution among flowering plants. In this study, we demonstrated chromosome behaviors in haploid B. rapa to understand how meiosis proceeds in presence of a single homolog. The findings showed that a diploid-like chromosome pairing was generally adapted during meiosis in haploid B. rapa. Non-homologous chromosomes in haploid cells paired at a high-frequency at metaphase I, over 50% of examined meiocytes showed at least three pairs of bivalents then equally segregated at anaphase I during meiosis. The fluorescence immunostaining showed that the cytoskeletal configurations were mostly well-organized during meiosis. Moreover, the expressed genes identified at meiosis in floral development was rather similar between haploid and diploid B. rapa, especially the expression of known hallmark genes pivotal to chromosome synapsis and homologous recombination were mostly in haploid B. rapa. Whole-genome duplication evolutionarily homology of genomic segments might be an important reason for this phenomenon, which would reshape the first division course of meiosis and influence pollen development in plants.

Two related families of metal transferases, ZNG1 and ZNG2, are involved in acclimation to poor Zn nutrition in Arabidopsis.

Metal homeostasis has evolved to tightly modulate the availability of metals within the cell, avoiding cytotoxic interactions due to excess and protein inactivity due to deficiency. Even in the presence of homeostatic processes, however, low bioavailability of these essential metal nutrients in soils can negatively impact crop health and yield. While research has largely focused on how plants assimilate metals, acclimation to metal-limited environments requires a suite of strategies that are not necessarily involved in metal transport across membranes. The identification of these mechanisms provides a new opportunity to improve metal-use efficiency and develop plant foodstuffs with increased concentrations of bioavailable metal nutrients. Here, we investigate the function of two distinct subfamilies of the nucleotide-dependent metallochaperones (NMCs), named ZNG1 and ZNG2, that are found in plants, using Arabidopsis thaliana as a reference organism. AtZNG1 (AT1G26520) is an ortholog of human and fungal ZNG1, and like its previously characterized eukaryotic relatives, localizes to the cytosol and physically interacts with methionine aminopeptidase type I (AtMAP1A). Analysis of AtZNG1, AtMAP1A, AtMAP2A, and AtMAP2B transgenic mutants are consistent with the role of Arabidopsis ZNG1 as a Zn transferase for AtMAP1A, as previously described in yeast and zebrafish. Structural modeling reveals a flexible cysteine-rich loop that we hypothesize enables direct transfer of Zn from AtZNG1 to AtMAP1A during GTP hydrolysis. Based on proteomics and transcriptomics, loss of this ancient and conserved mechanism has pleiotropic consequences impacting the expression of hundreds of genes, including those involved in photosynthesis and vesicle transport. Members of the plant-specific family of NMCs, ZNG2A1 (AT1G80480) and ZNG2A2 (AT1G15730), are also required during Zn deficiency, but their target protein(s) remain to be discovered. RNA-seq analyses reveal wide-ranging impacts across the cell when the genes encoding these plastid-localized NMCs are disrupted.

Comparative transcriptome analysis revealed differential gene expression in multiple signaling pathways at flowering in polyploid Brassica rapa.

BACKGROUND: Polyploidy is widespread in angiosperms and has a significant impact on plant evolution, diversity, and breeding program. However, the changes in the flower development regulatory mechanism in autotetraploid plants remains relatively limited. In this study, RNA-seq analysis was used to investigate changes in signaling pathways at flowering in autotetraploid Brassica rapa. RESULTS: The study findings showed that the key genes such as CO, CRY2, and FT which promotes floral formation were down-regulated, whereas floral transition genes FPF1 and FD were up-regulated in autotetraploid B. rapa. The data also demonstrated that the positive regulators GA1 and ELA1 in the gibberellin's biosynthesis pathway were negatively regulated by polyploidy in B. rapa. Furthermore, transcriptional factors (TFs) associated with flower development were significantly differentially expressed including the up-regulated CIB1 and AGL18, and the down-regulated AGL15 genes, and by working together such genes affected the expression of the down-stream flowering regulator FLOWERING LOCUS T in polyploid B. rapa. Compared with that in diploids autotetrapoid plants consist of differential expression within the signaling transduction pathway, with 13 TIFY gens up-regulated and 17 genes related to auxin pathway down-regulated. CONCLUSION: Therefore, polyploidy is more likely to integrate multiple signaling pathways to influence flowering in B. rapa after polyploidization. In general, the present results shed new light on our global understanding of flowering regulation in polyploid plants during breeding program.

Cytological and proteomic analyses of floral buds reveal an altered atlas of meiosis in autopolyploid Brassica rapa.

BACKGROUND: Polyploidy is considered as a basic event in plant speciation and evolution in nature, and the cytological and proteomic profilings of floral buds at meiosis (FAM) would definitely contribute to a better understanding of the polyploid-associated effects during plant reproduction cycle. RESULTS: Herein, the cytological investigations demonstrated that chromosome behaviors such as univalent and multivalent at prophase I, chaotic alignments at metaphase, aberrant segregation at telophase, were frequently observed during meiosis in autotetraploid Brassica rapa. The proteomic analysis showed a total of 562 differentially expressed proteins (DEPs) were identified in FAM between autotetraploid and diploid B. rapa. Notably, PARP2 and LIG1 related to base excision repair and BARD1 involved in recombination were significantly down-regulated in autotetraploid B. rapa, which indicated DNA repair pathway were more likely affected during meiosis in autotetraploid B. rapa. The functional analysis showed that DEPs assigned to "chromatin structure and dynamics", "cell cycle control, cell division, chromosome partitioning" and "cytoskeleton" were preferentially up-regulated, which suggested a robust regulation of cell division in autotetraploid B. rapa. In combination with the floral RNA-seq data released, a number of DEPs were found positively correlated with their transcript abundance, but posttranslational modification of proteins might also play a role in regulating meiosis course after polyploidization. CONCLUSIONS: In general, this study provides a detailed cytology and proteome landscape of FAM between diploid and autotetraploid B. rapa, which definitely affords us a better understanding of uniformity and discrepancy of meiosis at the plant reproductive stage before and after polyploidization.

Autopolyploidy leads to rapid genomic changes in Arabidopsis thaliana.

Polyploidy is a widespread feature of plant genomes. As a typical model of polyploidy, autopolyploidy has been postulated evolutionary dead ends and received little attention compared with allopolyploidy. For the limited data available so far, the evolutionary outcome of genome diversity in autopolyploids remains controversial in comparison with its diploid ancestors. In the present study, the effects of autopolyploidy on genome diversity were revealed at a genome-wide scale by comparative analyses of polymorphism between Arabidopsis autopolyploids (autotetraploids and autotriploids) and related diploids within the first ten successive inbred generations using amplified fragment length polymorphism. The results showed that in contrast with diploids, the rapid genomic changes (including gain and loss of DNA sequences) in autopolyploids were definitely found within the first generations after autopolyploidization, but slow down and probably stabilized in the higher generations as a source of genetic diversity in the long term. The sequencing of these DNA fragments indicated that these changes occurred both on genic and inter-genic (or intronic) regions, and quantitative PCR showed that the expression of some corresponding genes in the genic regions was obviously affected (including upregulation, downregulation and silencing) in autopolyploids. Therefore, this study demonstrated that autopolyploidy could lead to rapid genomic changes and probably influence expression and function of certain genes within the first generations, giving rising to genetic diversification after polyploidization.

Transcriptome Analysis of Floral Buds Deciphered an Irregular Course of Meiosis in Polyploid Brassica rapa.

Polyploidy is a fundamental process in plant evolution. Understanding the polyploidy-associated effects on plant reproduction is essential for polyploid breeding program. In the present study, our cytological analysis firstly demonstrated that an overall course of meiosis was apparently distorted in the synthetic polyploid Brassica rapa in comparison with its diploid progenitor. To elucidate genetic basis of this irregular meiosis at a molecular level, the comparative RNA-seq analysis was further used to investigate differential genetic regulation of developing floral buds identified at meiosis between autotetraploid and diploid B. rapa. In total, compared to its diploid counterparts, among all 40,927 expressed genes revealed, 4,601 differentially expressed genes (DEGs) were identified in the floral buds of autotetraploid B. rapa, among which 288 DEGs annotated were involved in meiosis. Notably, DMC1 identified as one previously known meiosis-specific gene involved in inter-homologous chromosome dependent repair of DNA double stranded breaks (DSBs), was significantly down-regulated in autotetraploid B. rapa, which presumably contributed to abnormal progression during meiosis I. Although certain DEGs associated with RNA helicase, cell cycling, and somatic DNA repair were up-regulated after genome duplication, genes associated with meiotic DSB repair were significantly down-regulated. Furthermore, the expression of randomly selected DEGs by RNA-seq analysis was confirmed by quantitative real-time PCR analysis in both B. rapa and Arabidopsis thaliana. Our results firstly account for adverse effects of polyploidy on an entire course of meiosis at both cytological and transcriptomic levels, and allow for a comprehensive understanding of the uniformity and differences in the transcriptome of floral buds at meiosis between diploid and polyploid B. rapa as well.