Jinfeng Zhao

DWARF3 Participates in an SCF Complex and Associates with DWARF14 to Suppress Rice Shoot Branching

Strigolactones (SLs) are a novel class of plant hormones that inhibit shoot branching. Currently, two proteins in rice are thought to play crucial roles in SL signal transduction. DWARF14 (D14), an α/β hydrolase, is responsible for SL perception, while DWARF3 (D3), an F-box protein with leucine-rich repeats, is essential for SL signal transduction. However, how these two proteins transmit SL signals to downstream factors remains unclear. Here, we characterized a high-tillering dwarf rice mutant, gsor300097, which is insensitive to GR24, a synthetic analog of SL. Mapping and sequencing analysis showed that gsor300097 is a novel allelic mutant of D3, in which a nonsense mutation truncates the protein from 720 to 527 amino acids. The D3 gene was strongly expressed in root, leaf, shoot base and panicle. Nuclear-localized F-box protein D3 played a role in the SCF complex by interacting with OSK1, OSK5 or OSK20 and OsCullin1. In addition, D3 associated with D14 in a GR24-dependent manner in vivo. Taken together, our findings suggested that D3 assembled into an SCF(D3) complex and associated with D14 to suppress rice shoot branching.

Characterization of a Null Allelic Mutant of the Rice NAL1 Gene Reveals Its Role in Regulating Cell Division

Leaf morphology is closely associated with cell division. In rice, mutations in Narrow leaf 1 (NAL1) show narrow leaf phenotypes. Previous studies have shown that NAL1 plays a role in regulating vein patterning and increasing grain yield in indica cultivars, but its role in leaf growth and development remains unknown. In this report, we characterized two allelic mutants of NARROW LEAF1 (NAL1), nal1-2 and nal1-3, both of which showed a 50% reduction in leaf width and length, as well as a dwarf culm. Longitudinal and transverse histological analyses of leaves and internodes revealed that cell division was suppressed in the anticlinal orientation but enhanced in the periclinal orientation in the mutants, while cell size remained unaltered. In addition to defects in cell proliferation, the mutants showed abnormal midrib in leaves. Map-based cloning revealed that nal1-2 is a null allelic mutant of NAL1 since both the whole promoter and a 404-bp fragment in the first exon of NAL1 were deleted, and that a 6-bp fragment was deleted in the mutant nal1-3. We demonstrated that NAL1 functions in the regulation of cell division as early as during leaf primordia initiation. The altered transcript level of G1- and S-phase-specific genes suggested that NAL1 affects cell cycle regulation. Heterogenous expression of NAL1 in fission yeast (Schizosaccharomyces pombe) further supported that NAL1 affects cell division. These results suggest that NAL1 controls leaf width and plant height through its effects on cell division.

Kinase activity of OsBRI1 is essential for brassinosteroids to regulate rice growth and development

Brassinosteroids (BRs) are steroid hormones that participate in multiple biological processes. In this paper, we characterized a classic rice mutant Fn189 (dwarf54, d54) showing semi-dwarf stature and erect leaves. The coleoptile elongation and root growth was less affected in Fn189 than wild-type plant by the exogenous application of eBL, the most active form of BRs. Lamina joint inclination assay and morphological analysis in darkness further showed that Fn189 mutant plant was insensitive to exogenous eBL. Through map-based cloning, Fn189 was found to be a novel allelic mutant of the DWARF 61 (D61) gene, which encodes the putative BRs receptor OsBRI1. A single base mutation caused the I834F substitution in the OsBRI1 kinase domain. Consequently, kinase activity of OsBRI1 was found to decrease dramatically. Taken together, the kinase activity of OsBRI1 is essential for brassinosteroids to regulate normal plant growth and development in rice.

A missense mutation in the transmembrane domain of CESA9 affects cell wall biosynthesis and plant growth in rice

Rice is a model organism in poaceae plants to study cell wall biosynthesis. In this study, a mutant S1-60 isolated from an EMS mutagenized japonica cultivar Nipponbare, is characterized by brittle culms that can be easily broken by bending. The reduction in mechanical strength was due to defect in thickening of the sclerenchyma cell wall. The amount of cellulose in S1-60 culms was reduced to 44.7% of that of wild-type plants. Besides, the mutant also exhibited pleiotropic phenotypes, such as dwarfism and partial sterility. Genetic analysis and map-based cloning showed that all the phenotype of S1-60 mutant was caused by a recessive point mutation in the OsCESA9 gene, which encodes the cellulose synthase A subunit 9. This yet uncharacterized missense mutation changed the highly conserved G905 to D at the beginning of the fifth transmembrane domain. The OsCESA9 gene is predominantly expressed in the culms of mature stage plants, consistent with the brittle phenotype in the culm. These results indicate that OsCESA9 plays an important role in cell wall biosynthesis and plant growth.

A Missense Mutation in the Zinc Finger Domain of OsCESA7 Deleteriously Affects Cellulose Biosynthesis and Plant Growth in Rice

Rice is a model plant species for the study of cellulose biosynthesis. We isolated a mutant, S1-24, from ethyl methanesulfonate (EMS)-treated plants of the japonica rice cultivar, Nipponbare. The mutant exhibited brittle culms and other pleiotropic phenotypes such as dwarfism and partial sterility. The brittle culms resulted from reduced mechanical strength due to a defect in thickening of the sclerenchyma cell wall and reduced cellulose content in the culms of the S1-24 mutant. Map-based gene cloning and a complementation assay showed that phenotypes of the S1-24 mutant were caused by a recessive point mutation in the OsCESA7 gene, which encodes cellulose synthase A subunit 7. The missense mutation changed the highly conserved C40 to Y in the zinc finger domain. The OsCESA7 gene is expressed predominantly in the culm at the mature stage, particularly in mechanical tissues such as vascular bundles and sclerenchyma cells, consistent with the brittle phenotype in the culm. These results indicate that OsCESA7 plays an important role in cellulose biosynthesis and plant growth.

Cloning and Expression of Gene Responsible for High-Tillering Dwarf Phenotype in Indica Rice Mutant gsor23

Abstract High-tillering dwarf mutant gsor23 was generated from an indica rice variety Indica9 radiatied by γ-ray. Genetic analysis showed that this phenotype was controlled by one single recessive gene, which was mapped within a physical distance of 386 kb between two insertion-deletion (InDel) markers C1-WT2 and C1-WT4 on the long arm of chromosome 1. There is a known gene D10 within this region, the mutation of which causes high-tillering in rice. Sequence analysis of the D10 allele in gsor23 revealed that the base cytosine (C) at the 404 th position in the coding region was deleted, which would cause frameshift mutation after the 134 th amino acids. The mutation site and indica background of gsor23 were different from the previously reported japonica mutants d10-1 and d10-2. Therefore, gsor23 is a novel allelic mutant of D10 which encodes the carotenoid-cleaving dioxygenase 8 (CCD8), a key enzyme involved in the biosynthesis of the new plant hormone strigolactones (SLs). After treatment with GR24, a synthetic analogue of SLs, the high-tillering phenotype of gsor23 was restored to normal. Real-time RT-PCR analysis showed that D10 expression was high in roots, but low in leaves. Compared with the wild type Indica9, the expression of the SL biosynthesis gene D10 was upregulated, while genes likely involved in the SL signal transduction pathway such as D3 and D14 were down-regulated in the gsor23 mutant.

The rice TRIANGULAR HULL1 protein acts as a transcriptional repressor in regulating lateral development of spikelet

As a basic unit of rice inflorescence, spikelet has profound influence on grain size, weight and yield. The molecular mechanism underlying spikelet development has not been fully elucidated. Here, we identified four allelic rice mutants, s2-89, xd151, xd281 and xd425, which exhibited reduced width of spikelet, especially in the apical region. Map-based cloning revealed that all these mutants had missense mutation in the TRIANGULAR HULL1 (TH1) gene, encoding an ALOG family protein. TH1 has been shown to regulate the lateral development of spikelet, but its mode of action remains unclear. Microscopic analysis revealed that the reduction in spikelet width was caused by decreased cell size rather than cell division. The TH1 protein was shown to localize in the nucleus and possess transcriptional repression activity. TH1 could form a homodimer and point mutation in the s2-89, xd281 and xd425 mutant inhibited homodimerization. The transcriptional repression activity of TH1 was partially relieved by the His129Tyr substitution in the s2-89 mutant. Fusion of an exogenous EAR transcription suppression domain to the mutant protein TH1s2-89 could largely complemented the narrow spikelet phenotype. These results indicate that TH1 functions as a transcription repressor and regulates cell expansion during the lateral development of spikelet.

Phenotypic and Genetic Analyses of a Novel Adaxially-Rolled Leaf Mutant in Rice

Leaf is an important organ for photosynthesis.Moderate rolling of leaves can facilitate the improvement of plants population structure and enhance light-use efficiency,which is very important in ideotype breeding of rice.In the present study,in order to systematically dissect the molecular mechanism of leaf morphogenesis and development,one ethyl methylsulfone(EMS)-induced rice(Oryza sativa L.)mutant with adaxially-rolled leaf,namely s1-145,was characterized.This mutant exhibited higher chlorophyll content,normal plant height and fertility.Genetic analysis indicated that the mutant was controlled by a single recessive gene.The mutated gene of s1-145 was fine mapped within a 90 kb interval between two InDel markers R2-34.70 and R2-34.79 on the long arm of chromosome 2 in rice.These results provide a basis for the final cloning and functional analysis of the leaf-rolling gene,as well as gene resource and plant material for rice ideotype breeding.