The 24-nucleotides (nt) phased secondary small interfering RNA (phasiRNA) is a unique class of plant small RNAs abundantly expressed in monocot anthers at early meiosis. Previously, 44 intergenic regions were identified as the loci for longer precursor RNAs of 24-nt phasiRNAs (24-PHASs) in the rice genome. However, the regulatory mechanism that determines spatiotemporal expression of these RNAs has remained elusive. ETERNAL TAPETUM1 (EAT1) is a basic-helix-loop-helix (bHLH) transcription factor indispensable for induction of programmed cell death (PCD) in postmeiotic anther tapetum, the somatic nursery for pollen production. In this study, EAT1-dependent non-cell-autonomous regulation of male meiosis was evidenced from microscopic observation of the eat1 mutant, in which meiosis with aberrantly decondensed chromosomes was retarded but accomplished somehow, eventually resulting in abortive microspores due to an aberrant tapetal PCD. EAT1 protein accumulated in tapetal-cell nuclei at early meiosis and postmeiotic microspore stages. Meiotic EAT1 promoted transcription of 24-PHAS RNAs at 101 loci, and importantly, also activated DICER-LIKE5 (DCL5, previous DCL3b in rice) mRNA transcription that is required for processing of double-stranded 24-PHASs into 24-nt lengths. From the results of the chromatin-immunoprecipitation and transient expression analyses, another tapetum-expressing bHLH protein, TDR INTERACTING PROTEIN2 (TIP2), was suggested to be involved in meiotic small-RNA biogenesis. The transient assay also demonstrated that UNDEVELOPED TAPETUM1 (UDT1)/bHLH164 is a potential interacting partner of both EAT1 and TIP2 during early meiosis. This study indicates that EAT1 is one of key regulators triggering meiotic phasiRNA biogenesis in anther tapetum, and that other bHLH proteins, TIP2 and UDT1, also play some important roles in this process. Spatiotemporal expression control of these bHLH proteins is a clue to orchestrate precise meiosis progression and subsequent pollen production non-cell-autonomously. Meiotic crossover formation shuffles homologous genes between parental genomes, and enables transmission of new gene sets to the offspring. Frequency and positions of crossovers are determined by numerous genetic and epigenetic factors, and low nucleosome-density regions are associated with crossover hot spots in yeasts and Arabidopsis. The epigenetic chromosome landscape is shaped by unevenly distributed modifications of nucleosome components, histones and DNAs. Recently, we found that MEL1 (ARGONAUTE5) promotes large-scale remodeling of meiotic chromosomes with dramatic increases of histone H3 lysine 9 dimethylation, and that loss of MEL1 resulted in early meiotic arrest with few crossovers present. In rice anthers, MEL1-associating small interfering RNAs (masiRNAs) were composed of large amounts of premeiotic 21-nt phasiRNAs, plus low levels of both 24-nt repeat-associated siRNA and meiotic 24-nt phasiRNAs. Production of 24-nt phasiRNA during the meiotic stage was largely EAT1-dependent. Collectively, our findings suggest a possibility that unknown small RNA-mediated signaling regulates male meiosis non-cell-autonomously, probably a downstream output involves large-scale chromosome remodeling promoted by Argonaute proteins, while a possibility of EAT1-dependent, but small RNA-independent signaling cannot be excluded. In any cases, the studies on MEL1 and tapetal bHLH proteins will be a clue to reveal small RNA-mediated processes determining meiotic epigenetic landscape.

The roles of epigenetic mechanisms, including small-RNA-mediated silencing, in plant meiosis largely remain unclear, despite their importance in plant reproduction. This study unveiled that rice chromosomes are reprogrammed during the premeiosis-to-meiosis transition in pollen mother cells (PMCs). This large-scale meiotic chromosome reprogramming (LMR) continued throughout meiosis I, during which time H3K9 dimethylation (H3K9me2) was increased, and H3K9 acetylation and H3S10 phosphorylation were broadly decreased, with an accompanying immunostaining pattern shift of RNA polymerase II. LMR was dependent on the rice Argonaute protein, MEIOSIS ARRESTED AT LEPTOTENE1 (MEL1), which is specifically expressed in germ cells prior to meiosis, because LMR was severely diminished in mel1 mutant anthers. Pivotal meiotic events, such as pre-synaptic centromere association, DNA double-strand break initiation and synapsis of homologous chromosomes, were also disrupted in this mutant. Interestingly, and as opposed to the LMR loss in most chromosomal regions, aberrant meiotic protein loading and hypermethylation of H3K9 emerged on the nucleolar organizing region in the mel1 PMCs. These results suggest that MEL1 plays important roles in epigenetic LMR to promote faithful homologous recombination and synapsis during rice meiosis.

Salt spray is a major environmental issue in coastal areas. Cinnamomum camphora is an economically important tree species that grows in the coastal areas of southern China. Arbuscular mycorrhizal fungi (AMF) can alleviate the detrimental effects of abiotic stress on host plants. However, the mechanism by which AMF mitigates the adverse effects of salt spray on C. camphora remains unclear. A pot experiment was conducted in a greenhouse, where C. camphora seedlings were exposed to four AMF regimes (inoculation with sterilized fungi, with Glomus tortuosum, Funneliformis mosseae, either alone or in combination) and three salt spray regimes (applied with distilled water, 7, and 14 mg NaCl cm⁻²) in order to investigate the influence on root functional traits and plant growth. The results showed that higher salt spray significantly decreased the K⁺ uptake, K⁺/Na⁺ ratio, N/P ratio, total dry weight, and salinity tolerance of non-mycorrhizal plants by 37.9%, 71%, 27.4%, 12.7%, and 221.3%, respectively, when compared with control plants grown under non-salinity conditions. Mycorrhizal inoculation, particularly with a combination of G. tortuosum and F. mosseae, greatly improved the P uptake, total dry weight, and salinity tolerance of plants grown under higher salt spray conditions by 51.0%, 36.7%, and 130.9%, respectively, when compared with their counterparts. The results show that AMF can alleviate the detrimental effects of salt spray on C. camphora seedlings. Moreover, an enhanced uptake of K⁺ and P accounted for the resistance of the plants to salt spray. Therefore, pre-inoculation with a combination of G. tortuosum and F. mosseae to improve nutrient acquisition is a potential method of protecting C. camphora plants against salt spray stress in coastal areas.

The development of floral organs is coordinated by an elaborate network of homeotic genes, and gibberellin (GA) signaling is involved in floral organ development; however, the underlying molecular mechanisms remain elusive. In the present study, we found that MOS4-ASSOCIATED COMPLEX 5A (MAC5A), which is a protein containing an RNA-binding motif, was involved in the development of sepals, petals, and stamens; either the loss or gain of MAC5A function resulted in stamen malformation and a reduced seed set. The exogenous application of GA considerably exacerbated the defects in mac5a null mutants, including fewer stamens and male sterility. MAC5A was predominantly expressed in pollen grains and stamens, and overexpression of MAC5A affected the expression of homeotic genes such as APETALA1 (AP1), AP2, and AGAMOUS (AG). MAC5A may interact with RABBIT EARS (RBE), a repressor of AG expression in Arabidopsis flowers. The petal defect in rbe null mutants was at least partly rescued in mac5a rbe double mutants. These findings suggest that MAC5A is a novel factor that is required for the normal development of stamens and depends on the GA signaling pathway.