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The mechanisms underlying the photo-ammonification of nitrogenous organic compounds (NOCs) remain unclear, partly due to the analytical challenges of small NOC intermediates. This study introduced a simple methodology for accurately and simultaneously quantifying multiple small NOCs during ammonification processes. The developed method employed phenyl isothiocyanate as derivatization reagents, followed by high-performance liquid chromatography analysis to measure primary and secondary amines, amides, as well as NH4+ over variable photo-ammonification conditions. In our experimental setup, vacuum ultraviolet (VUV) irradiation serves as the controlled reaction environment to simulate harsh photo-ammonification environment. Representative NOCs, including pyridine, N,N-dimethylformamide, and acrylonitrile, were chosen due to their structural diversity and environmental relevance as model NOCs. This method was able to achieve excellent nitrogen mass balance, and revealed that the last steps of photo-ammonification involved oxidation of nitrogen-adjacent carbon to amide followed by the cleavage of N–C bond. This novel method may also help quantitative investigation of nitrogen transformations in different environmental contexts. [ABSTRACT FROM AUTHOR]

Background: As an important subfamily of arabinogalactan proteins (AGPs), fasciclin-like AGPs (FLAs) contribute to various aspects of growth, development and adaptation, yet their function remains largely elusive. Despite the diversity of FLAs, only two members, Arabidopsis FLA3 and rice MTR1, are reported to be involved in sexual reproduction. In this study, another Arabidopsis FLA-encoding gene, FLA14, was identified, and its role was investigated. Results: Arabidopsis FLA14 was found to be a pollen grain-specific gene. Expression results from fusion with green fluorescent protein showed that FLA14 was localized along the cell membrane and in Hechtian strands. A loss-of-function mutant of FLA14 showed no discernible defects during male gametogenesis, but precocious pollen germination occurred inside the mature anthers under high moisture conditions. Overexpression of FLA14 caused 39.2% abnormal pollen grains with a shrunken and withered appearance, leading to largely reduced fertility with short mature siliques and lower seed set. Cytological and ultramicroscopic observation showed that ectopic expression of FLA14 caused disruption at the uninucleate stage, resulting in either collapsed pollen with absent intine or pollen of normal appearance but with a thickened intine. Conclusions: Taken together, our data suggest a role for FLA14 in pollen development and preventing premature pollen germination inside the anthers under high relative humidity in Arabidopsis. [ABSTRACT FROM AUTHOR]

Efficient mRNA splicing is a prerequisite for protein biosynthesis and the eukaryotic splicing machinery is evolutionarily conserved among species of various phyla. At its catalytic core resides the activated splicing complex Bact consisting of the three small nuclear ribonucleoprotein complexes (snRNPs) U2, U5 and U6 and the so-called NineTeen complex (NTC) which is important for spliceosomal activation. CWC15 is an integral part of the NTC in humans and it is associated with the NTC in other species. Here we show the ubiquitous expression and developmental importance of the Arabidopsis ortholog of yeast CWC15. CWC15 associates with core components of the Arabidopsis NTC and its loss leads to inefficient splicing. Consistent with the central role of CWC15 in RNA splicing, cwc15 mutants are embryo lethal and additionally display strong defects in the female haploid phase. Interestingly, the haploid male gametophyte or pollen in Arabidopsis, on the other hand, can cope without functional CWC15, suggesting that developing pollen might be more tolerant to CWC15-mediated defects in splicing than either embryo or female gametophyte. [ABSTRACT FROM AUTHOR]

Ogura-type cytoplasmic male sterility (Ogura-CMS) has been widely used in the hybrid breeding industry for cruciferous vegetables. Turnip (Brassica rapa ssp. rapifera) is one of the most important local cruciferous vegetables in China, cultivated for its fleshy root as a flat disc. Here, morphological characteristics of an Ogura-CMS line ‘BY10-2A’ and its maintainer fertile (MF) line ‘BY10-2B’ of turnip were investigated. Ogura-CMS turnip showed a reduction in the size of the fleshy root, and had distinct defects in microspore development and tapetum degeneration during the transition from microspore mother cells to tetrads. Defective microspore production and premature tapetum degeneration during microgametogenesis resulted in short filaments and withered white anthers, leading to complete male sterility of the Ogura-CMS line. Additionally, the mechanism regulating Ogura-CMS in turnip was investigated using inflorescence transcriptome analyses of the Ogura-CMS and MF lines. The de novo assembly resulted in a total of 84,132 unigenes. Among them, 5,117 differentially expressed genes (DEGs) were identified, including 1,339 up- and 3,778 down-regulated genes in the Ogura-CMS line compared to the MF line. A number of functionally known members involved in anther development and microspore formation were addressed in our DEG pool, particularly genes regulating tapetum programmed cell death (PCD), and associated with pollen wall formation. Additionally, 185 novel genes were proposed to function in male organ development based on GO analyses, of which 26 DEGs were genotype-specifically expressed. Our research provides a comprehensive foundation for understanding anther development and the CMS mechanism in turnip. [ABSTRACT FROM AUTHOR]

Summary: Arabinogalactan proteins (AGPs) are extensively glycosylated hydroxyproline‐rich glycoproteins ubiquitous in all plant tissues and cells. AtAGP6 and AtAGP11, the only two functionally known pollen‐specific classical AGP encoding genes in Arabidopsis, are reported to have redundant functions in microspore development. BcMF18 and BcMF8 isolated from Brassica campestris are the orthologues of AtAGP6 and AtAGP11, respectively. In contrast to the functional redundancy of AtAGP6 and AtAGP11, single‐gene disruption of BcMF8 led to deformed pollen grains with abnormal intine development and ectopic aperture formation in B. campestris. Here, we further explored the action of BcMF18 and its relationship with BcMF8. BcMF18 was specifically expressed in pollen during the late stages of microspore development. Antisense RNA transgenic lines with BcMF18 reduction resulted in aberrant pollen grains with abnormal cellulose distribution, lacking intine, cytoplasm and nuclei. Transgenic plants with repressive expression of both BcMF8 and BcMF18 showed a hybrid phenotype, expressing a mixture of the phenotypes of the single gene knockdown plant lines. In addition, we identified functional diversity between BcMF18/BcMF8 and AtAGP6/AtAGP11, mainly reflected by the specific contribution of BcMF18 and BcMF8 to pollen wall formation. These results suggest that, unlike the orthologous genes AtAGP6 and AtAGP11 in Arabidopsis, BcMF18 and BcMF8 are both integral to pollen biogenesis in B. campestris, acting through independent pathways during microspore development. [ABSTRACT FROM AUTHOR]

Arabinogalactan proteins (AGPs) are a superfamily of hydroxyproline-rich glycoproteins that are massively glycosylated, widely implicated in plant growth and development. No comprehensive analysis of the AGP gene family has been performed in Chinese cabbage (Brassica rapa ssp. chinensis). Here, we identified a total of 293 putative AGP-encoding genes in B. rapa, including 25 classical AGPs, three lysine-rich AGPs, 30 AG-peptides, 36 fasciclin-like AGPs (FLAs), 59 phytocyanin-like AGPs, 33 xylogen-like AGPs, 102 other chimeric AGPs, two non-classical AGPs and three AGP/extensin hybrids. Their protein structures, phylogenetic relationships, chromosomal location and gene duplication status were comprehensively analyzed. Based on RNA sequencing data, we found that 73 AGP genes were differentially expressed in the floral buds of the sterile and fertile plants at least at one developmental stage in B. rapa, suggesting a potential role of AGPs in male reproductive development. We further characterized BrFLA2, BrFLA28 and BrFLA32, three FLA members especially expressed in anthers, pollen grains and pollen tubes. BrFLA2, BrFLA28 and BrFLA32 are indispensable for the proper timing of pollen germination under high relative humidity. Our study greatly extends the repertoire of AGPs in B. rapa and reveals a role for three members of the FLA subfamily in pollen germination. [ABSTRACT FROM AUTHOR]