Jing-Liu Zhang

Over-expression of rice OsAGO7 gene induces upward curling of the leaf blade that enhanced erect-leaf habit

High-yield cultivars are characterized by erect leaf canopies that optimize photosynthesis and thus favor increased biomass. Upward curling of the leaf blade (called rolled leaf) can result in enhanced erect-leaf habit, increase erect duration and promote an overall erect leaf canopy. The rice mutant R05, induced through transferred DNA (T-DNA) insertion, had the rolled-leaf trait. The leaves in the wild type demonstrated natural drooping tendencies, resulting in decreasing leaf erection indices (LEIs) during senescence at the 20th day after flowering. Conversely, LEIs of the leaves in R05 remained high, even 20-day post-flowering. We applied T-DNA tagging and isolated a rolled-leaf gene from rice which, when over-expressed, could induce upward curling of the leaf blade. This gene encodes for a protein of 1,048 amino acids including the PAZ and PIWI conserved domains, belonging to the Argonaute (AGO) family. There are at least 18 members of the AGO family in rice. According to high-sequence conservation, the rolled-leaf gene in rice could be orthologous to the Arabidopsis ZIP/Ago7 gene, so we called it OsAGO7. These results provide a possible opportunity for implementing OsAGO7 gene in crop improvement.

Overexpression of ACL1 (abaxially curled leaf 1) Increased Bulliform Cells and Induced Abaxial Curling of Leaf Blades in Rice

Understanding the genetic mechanism underlying rice leaf-shape development is crucial for optimizing rice configuration and achieving high yields; however, little is known about leaf abaxial curling. We isolated a rice transferred DNA (T-DNA) insertion mutant, BY240, which exhibited an abaxial leaf curling phenotype that co-segregated with the inserted T-DNA. The T-DNA was inserted in the promoter of a novel gene, ACL1 (Abaxially Curled Leaf 1), and led to overexpression of this gene in BY240. Overexpression of ACL1 in wild-type rice also resulted in abaxial leaf curling. ACL1 encodes a protein of 116 amino acids with no known conserved functional domains. Overexpression of ACL2, the only homolog of ACL1 in rice, also induced abaxial leaf curling. RT-PCR analysis revealed high expressions of ACLs in leaf sheaths and leaf blades, suggesting a role for these genes in leaf development. In situ hybridization revealed non-tissue-specific expression of the ACLs in the shoot apical meristem, leaf primordium, and young leaf. Histological analysis showed increased number and exaggeration of bulliform cells and expansion of epidermal cells in the leaves of BY240, which caused developmental discoordination of the abaxial and adaxial sides, resulting in abaxially curled leaves. These results revealed an important mechanism in rice leaf development and provided the genetic basis for agricultural improvement.

Characterization of Rolled and Erect Leaf 1 in regulating leave morphology in rice

Highlight The Rolled and Erect Leaf 1 (REL1) gene is a novel component controlling brassinosteroid signalling-associated leaf morphogenesis and leaf angle in Oryza sativa.

Overexpression of Osta-siR2141 caused abnormal polarity establishment and retarded growth in rice

Small RNAs (smRNAs) including miRNAs and siRNAs are critical for gene regulation and plant development. Among the highly diverse siRNAs, trans-acting siRNAs (ta-siRNAs) have been shown to be plant-specific. In Arabidopsis, eight TAS loci belonging to four families (TAS1, TAS2, TAS3, and TAS4) have been identified, and bioinformatics analysis reveals that the sequence of TAS3 is highly conserved in plants. In this study, the function of TAS3 ta-siRNA (tasiR-ARF) has been revealed in rice (Oryza sativa L.) on polarity establishment and stage transition from vegetative to reproductive development by over-expressing Osta-siR2141. Osta-siR2141 replaced miR390 in the miR390 backbone for ectopic expression in rice, and overexpression of Osta-siR2141 caused disturbed vascular bundle development and adaxialization in polarity establishment. Transgenic lines also displayed abnormal shoot apical meristems (SAMs) and retarded growth at the vegetative stage. Molecular analysis revealed that overexpression of Osta-siR2141 resulted in the down-regulation of miR166 and the up-regulation of class III homeodomain-leucine zipper genes (HD-ZIPIIIs) in the vegetative stage but not in the reproductive stage. Moreover, overexpression of Osta-siR2141 in Arabidopsis disturbed polarity establishment and retarded stage transition, suggesting that tasiR-ARF was functionally conserved in rice and Arabidopsis.

An atypical HLH protein OsLF in rice regulates flowering time and interacts with OsPIL13 and OsPIL15.

In plants, flowering as a crucial developmental event is highly regulated by both genetic programs and environmental signals. Genetic analysis of flowering time mutants is instrumental in dissecting the regulatory pathways of flowering induction. In this study, we isolated the OsLF gene by its association with the T-DNA insertion in the rice late flowering mutant named A654. The OsLF gene encodes an atypical HLH protein composed of 419 amino acids (aa). Overexpression of the OsLF gene in wild type rice recapitulated the late flowering phenotype of A654, indicating that the OsLF gene negatively regulates flowering. Flowering genes downstream of OsPRR1 such as OsGI and Hd1 were down regulated in the A654 mutant. Yeast two hybrid and colocalization assays revealed that OsLF interacts strongly with OsPIL13 and OsPIL15 that are potentially involved in light signaling. In addition, OsPIL13 and OsPIL15 colocalize with OsPRR1, an ortholog of the Arabidopsis APRR1 gene that controls photoperiodic flowering response through clock function. Together, these results suggest that overexpression of OsLF might repress expression of OsGI and Hd1 by competing with OsPRR1 in interacting with OsPIL13 and OsPIL15 and thus induce late flowering.

Albino Leaf1 That Encodes the Sole Octotricopeptide Repeat Protein Is Responsible for Chloroplast Development

An octotricopeptide repeat protein regulates chloroplast development by coordinating the transcription and translation of chloroplast associated genes in rice. Chloroplast, the photosynthetic organelle in plants, plays a crucial role in plant development and growth through manipulating the capacity of photosynthesis. However, the regulatory mechanism of chloroplast development still remains elusive. Here, we characterized a mutant with defective chloroplasts in rice (Oryza sativa), termed albino leaf1 (al1), which exhibits a distinct albino phenotype in leaves, eventually leading to al1 seedling lethality. Electronic microscopy observation demonstrated that the number of thylakoids was reduced and the structure of thylakoids was disrupted in the al1 mutant during rice development, which eventually led to the breakdown of chloroplast. Molecular cloning revealed that AL1 encodes the sole octotricopeptide repeat protein (RAP) in rice. Genetic complementation of Arabidopsis (Arabidopsis thaliana) rap mutants indicated that the AL1 protein is a functional RAP. Further analysis illustrated that three transcript variants were present in the AL1 gene, and the altered splices occurred at the 3′ untranslated region of the AL1 transcript. In addition, our results also indicate that disruption of the AL1 gene results in an altered expression of chloroplast-associated genes. Consistently, proteomic analysis demonstrated that the abundance of photosynthesis-associated proteins is altered significantly, as is that of a group of metabolism-associated proteins. More specifically, we found that the loss of AL1 resulted in altered abundances of ribosomal proteins, suggesting that RAP likely also regulates the homeostasis of ribosomal proteins in rice in addition to the ribosomal RNA. Taken together, we propose that AL1, particularly the AL1a and AL1c isoforms, plays an essential role in chloroplast development in rice.

Dense-panicle-related gene cloning from rice mutant A989 and transgenic plant analysis.

Abstract Flowering time and inflorescence architecture are important and inter-relating traits in rice (Oryza sativa L.). Overexpression of each of RCN1 to RCN3 in rice could result in delayed flowering and abnormal inflorescence architecture. From a transferred DNA (T-DNA) insertion population constructed from japonica rice cv. Zhonghua 11, a mutant, A989, exhibiting dense panicle and late flowering was isolated. Genetic and molecular analysis showed that a single copy of T-DNA was inserted into the genome of the mutant, and the T-DNA was inserted into the 330 bp position after RCN2 poly A. Overexpression of RCN2 was observed in the mutant as revealed by reverse transcription-polymerase chain reaction (RT-PCR) test. When RCN2 gene was transferred into wild type driven by double 35S promoters, the transformants showed no panicle due to the suspensions of differentiation and growth of the secondary-branch primordium; however, the transition from vegetative growth to productive growth was normal in the transgenic plant.