Effect of Panicle Morphology on Grain Filling and Rice Yield: Genetic Control and Molecular Regulation.

The demand for rice is likely to increase approximately 1.5 times by the year 2050. In contrast, the rice production is stagnant since the past decade as the ongoing rice breeding program is unable to increase the production further, primarily because of the problem in grain filling. Investigations have revealed several reasons for poor filling of the grains in the inferior spikelets of the compact panicle, which are otherwise genetically competent to develop into well-filled grains. Among these, the important reasons are 1) poor activities of the starch biosynthesizing enzymes, 2) high ethylene production leading to inhibition in expressions of the starch biosynthesizing enzymes, 3) insufficient division of the endosperm cells and endoreduplication of their nuclei, 4) low accumulation of cytokinins and indole-3-acetic acid (IAA) that promote grain filling, and 5) altered expressions of the miRNAs unfavorable for grain filling. At the genetic level, several genes/QTLs linked to the yield traits have been identified, but the information so far has not been put into perspective toward increasing the rice production. Keeping in view the genetic competency of the inferior spikelets to develop into well-filled grains and based on the findings from the recent research studies, improving grain filling in these spikelets seems plausible through the following biotechnological interventions: 1) spikelet-specific knockdown of the genes involved in ethylene synthesis and overexpression of ß-CAS (ß-cyanoalanine) for enhanced scavenging of CN- formed as a byproduct of ethylene biosynthesis; 2) designing molecular means for increased accumulation of cytokinins, abscisic acid (ABA), and IAA in the caryopses; 3) manipulation of expression of the transcription factors like MYC and OsbZIP58 to drive the expression of the starch biosynthesizing enzymes; 4) spikelet-specific overexpression of the cyclins like CycB;1 and CycH;1 for promoting endosperm cell division; and 5) the targeted increase in accumulation of ABA in the straw during the grain filling stage for increased carbon resource remobilization to the grains. Identification of genes determining panicle compactness could also lead to an increase in rice yield through conversion of a compact-panicle into a lax/open one. These efforts have the ability to increase rice production by as much as 30%, which could be more than the set production target by the year 2050.

Genes determining panicle morphology and grain quality in rice (Oryza sativa).

The world's increase in rice (Oryza sativa L.) production is not keeping up with the increase in its population. To boost the introduction of new high-yielding cultivars, knowledge is being gained on the genes and quantitative trait loci (QTLs) determining the panicle phenotype. The important are those determining yield of the crop, such as grain numbers per panicle and size and weight of the grains. Biochemical and molecular functions of many of them are understood in some details. Among these, OsCKX2 and OsSPL14 have been shown to increase panicle branching and grain numbers when overexpressed. Furthermore, miRNAs appear to play an important role in determining the panicle morphology by regulating the expressions of the genes like OsSPL14 and GRF4 involved in panicle branching and grain numbers and length. Mutations also greatly influence the grain shape and size. However, the information gained so far on the genetic regulation of grain filling and panicle morphology has not been successfully put into commercial application. Furthermore, the identification of the gene(s)/QTLs regulating panicle compactness is still lacking, which may enable the researchers to convert a compact-panicle cultivar into a lax/open one, and thereby increasing the chances of enhancing the yield of a desired compact-panicle cultivar obtained by the breeding effort.

Biochemical and molecular processes contributing to grain filling and yield in rice.

The increase in much required rice production through breeding programmes is on decline. The primary reason being poor filling of grains in the basal spikelets of the heavy and compact panicle rice developed. These spikelets are genetically competent to develop into well filled grains, but fail to do so because the carbohydrate assimilates available to them remain unutilized, reportedly due to poor activities of the starch biosynthesizing enzymes, high production of ethylene leading to enhanced synthesis of the downstream signaling component RSR1 protein that inhibits GBSS1 activity, poor endosperm cell division and endoreduplication of the endosperm nuclei, altered expression of the transcription factors influencing grain filling, enhanced expression and phosphorylation of 14-3-3 proteins, poor expression of the seed storage proteins, reduced synthesis of the hormones like cytokinins and IAA that promote grain filling, and altered expression of miRNAs preventing their normal role in grain filling. Since the basal spikelets are genetically competent to develop into well filled mature grains, biotechnological interventions in terms of spikelet-specific overexpression of the genes encoding enzymes involved in grain filling and/or knockdown/overexpression of the genes influencing the activities of the starch biosynthesizing enzymes, various cell cycle events and hormone biosynthesis could increase rice production by as much as 30%, much more than the set production target of 800 mmt. Application of these biotechnological interventions in the heavy and compact panicle cultivars producing grains of desired quality would also maintain the quality of the grains having demand in market besides increasing the rice production per se.

Study of expressions of miRNAs in the spikelets based on their spatial location on panicle in rice cultivars provided insight into their influence on grain development.

Development of rice cultivars bearing numerous spikelets by breeding approach to increase the yearly production of rice to approximately 800 million metric tons to feed the ever increasing population of the world accompanies poor grain filling in the inferior spikelets preventing achievement of the yield potential. As the initial stages of caryopses development are of much importance for grain filling, spatio-temporal expressions of the miRNAs were studied during these periods in the spikelets of a compact-panicle rice cultivar, Oryza sativa cv. Mahalaxmi, bearing numerous spikelets per panicle to understand the reason of poor grain filling at the level of the initial biochemical events. Differential expression of several known miRNAs between the superior and inferior spikelets suggested great difference in metabolism related to grain filling in the spikelets based on their spatial location on compact panicle. Expressions of five known and four novel miRNAs were validated by Northern. Their targets included the enzymes directly involved in starch biosynthesis like sucrose synthase, starch synthase and pullulanase, besides others. Spatio-temporal expression studies of these miRNAs in the spikelets of Mahalaxmi revealed a pattern of mostly a greater expression in the inferior spikelets compared with the superior ones concomitant with an inverse expression of the target genes, which was not observed in the lax-panicle cultivar Upahar. The study thus revealed that the grain filling in rice is greatly regulated by miRNAs, and these miRNAs or their target genes could be considered for biotechnological interventions for improving grain filling in the rice cultivars of interest.

Biochemical and molecular characterisation of exogenous cytokinin application on grain filling in rice.

BACKGROUND: Poor filling of grains in the basal spikelets of large size panicles bearing numerous spikelets has been a major limitation in attempts to increase the rice production to feed the world's increasing population. Considering that biotechnological intervention could play important role in overcoming this limitation, the role of cytokinin in grain filling was investigated based on the information on cell proliferating potential of the hormone and reports of its high accumulation in immature seeds. RESULTS: A comparative study considering two rice varieties differing in panicle compactness, lax-panicle Upahar and compact-panicle OR-1918, revealed significant difference in grain filling, cytokinin oxidase (CKX) activity and expression, and expression of cell cycle regulators and cytokinin signaling components between the basal and apical spikelets of OR-1918, but not of Upahar. Exogenous application of cytokinin (6-Benzylaminopurine, BAP) to OR-1918 improved grain filling significantly, and this was accompanied by a significant decrease in expression and activity of CKX, particularly in the basal spikelets where the activity of CKX was significantly higher than that in the apical spikelets. Cytokinin application also resulted in significant increase in expression of cell cycle regulators like cyclin dependent kinases and cyclins in the basal spikelets that might be facilitating cell division in the endosperm cells by promoting G1/S phase and G2/M phase transition leading to improvement in grain filling. Expression studies of type-A response regulator (RR) component of cytokinin signaling indicated possible role of OsRR3, OsRR4 and OsRR6 as repressors of CKX expression, much needed for an increased accumulation of CK in cells. Furthermore, the observed effect of BAP might not be solely because of it, but also because of induced synthesis of trans-zeatin (tZ) and N6-(Δ2-isopentenyl)adenine (iP), as reflected from accumulation of tZR (tZ riboside) and iPR (iP riboside), and significantly enhanced expression of an isopentenyl transferase (IPT) isoform. CONCLUSION: The results suggested that seed-specific overexpression of OsRR4 and OsRR6, and more importantly of IPT9 could be an effective biotechnological intervention towards improving the CK level of the developing caryopses leading to enhanced grain filling in rice cultivars bearing large panicles with numerous spikelets, and thereby increasing their yield potential.

Identification and Characterization of Differentially Expressed Genes in Inferior and Superior Spikelets of Rice Cultivars with Contrasting Panicle-Compactness and Grain-Filling Properties.

Breeding programs for increasing spikelet number in rice have resulted in compactness of the panicle, accompanied by poor grain filling in inferior spikelets. Although the inefficient utilization of assimilate has been indicated as responsible for this poor grain filling, the underlying cause remains elusive. The current study utilized the suppression subtractive hybridization technique to identify 57 and 79 genes that overexpressed in the superior and inferior spikelets (with respect to each other), respectively, of the compact-panicle rice cultivar Mahalaxmi. Functional categorization of these differentially expressed genes revealed a marked metabolic difference between the spikelets according to their spatial location on the panicle. The expression of genes encoding seed storage proteins was dominant in inferior spikelets, whereas genes encoding regulatory proteins, such as serine-threonine kinase, zinc finger protein and E3 ligase, were highly expressed in superior spikelets. The expression patterns of these genes in the inferior and superior spikelets of Mahalaxmi were similar to those observed in another compact-panicle cultivar, OR-1918, but differed from those obtained in two lax-panicle cultivars, Upahar and Lalat. The results first suggest that the regulatory proteins abundantly expressed in the superior spikelets of compact-panicle cultivars and in both the superior and inferior spikelets of lax-panicle cultivars but poorly expressed in the inferior spikelets of compact-panicle cultivars promote grain filling. Second, the high expression of seed-storage proteins observed in the inferior spikelets of compact-panicle cultivars appears to inhibit the grain filling process. Third, the low expression of enzymes of the Krebs cycle in inferior spikelets compared with superior spikelets of compact-panicle cultivars is bound to lead to poor ATP generation in the former and consequently limit starch biosynthesis, an ATP-consuming process, resulting in poor grain filling.