Summary
The CRISPR/Cas9 system has been demonstrated to efficiently induce targeted gene editing in a variety of organisms including plants. Recent work showed that CRISPR/Cas9‐induced gene mutations ...in Arabidopsis were mostly somatic mutations in the early generation, although some mutations could be stably inherited in later generations. However, it remains unclear whether this system will work similarly in crops such as rice. In this study, we tested in two rice subspecies 11 target genes for their amenability to CRISPR/Cas9‐induced editing and determined the patterns, specificity and heritability of the gene modifications. Analysis of the genotypes and frequency of edited genes in the first generation of transformed plants (T0) showed that the CRISPR/Cas9 system was highly efficient in rice, with target genes edited in nearly half of the transformed embryogenic cells before their first cell division. Homozygotes of edited target genes were readily found in T0 plants. The gene mutations were passed to the next generation (T1) following classic Mendelian law, without any detectable new mutation or reversion. Even with extensive searches including whole genome resequencing, we could not find any evidence of large‐scale off‐targeting in rice for any of the many targets tested in this study. By specifically sequencing the putative off‐target sites of a large number of T0 plants, low‐frequency mutations were found in only one off‐target site where the sequence had 1‐bp difference from the intended target. Overall, the data in this study point to the CRISPR/Cas9 system being a powerful tool in crop genome engineering.
Abscisic acid (ABA) is an important phytohormone regulating plant growth, development, and stress responses. It has an essential role in multiple physiological processes of plants, such as stomatal ...closure, cuticular wax accumulation, leaf senescence, bud dormancy, seed germination, osmotic regulation, and growth inhibition among many others. Abscisic acid controls downstream responses to abiotic and biotic environmental changes through both transcriptional and posttranscriptional mechanisms. During the past 20 years, ABA biosynthesis and many of its signaling pathways have been well characterized. Here we review the dynamics of ABA metabolic pools and signaling that affects many of its physiological functions.
Abscisic acid (ABA) is the major stress hormone that coordinates plant growth, development and abiotic stress responses. In this review, we summarized the recent progresses on its metabolism, transport and signaling, and discussed the open questions about ABA dynamics and functions.
Domestication has resulted in reduced salt tolerance in tomato. To identify the genetic components causing this deficiency, we performed a genome‐wide association study (GWAS) for root Na+/K+ ratio ...in a population consisting of 369 tomato accessions with large natural variations. The most significant variations associated with root Na+/K+ ratio were identified within the gene SlHAK20 encoding a member of the clade IV HAK/KUP/KT transporters. We further found that SlHAK20 transports Na+ and K+ and regulates Na+ and K+ homeostasis under salt stress conditions. A variation in the coding sequence of SlHAK20 was found to be the causative variant associated with Na+/K+ ratio and confer salt tolerance in tomato. Knockout mutations in tomato SlHAK20 and the rice homologous genes resulted in hypersensitivity to salt stress. Together, our study uncovered a previously unknown molecular mechanism of salt tolerance responsible for the deficiency in salt tolerance in cultivated tomato varieties. Our findings provide critical information for molecular breeding to improve salt tolerance in tomato and other crops.
Synopsis
Selection of large fruits in domesticated tomato is linked to a reduction in salt tolerance. This study links domestication‐associated variation in the Na+/K+ transporter‐coding gene SlHAK20 to reduced salt tolerance in cultivated plants.
Root Na+/K+ ratios show a strong positive correlation with fruit weight during tomato domestication.
A variation in SlHAK20 is associated with Na+/K+ ratio modulation in tomato roots under salt stress conditions.
The SlHAK20Hap1 haplotype is more effective than SlHAK20Hap2 in conferring Na+ homeostasis and salt tolerance.
The role of SlHAK20 in regulation of salt tolerance is conserved in rice.
A genome‐wide association study links reduced sodium ion uptake by SlHAK20 transporter to diminished salt tolerance and increased fruit weight in cultivated tomato.
A major problem facing humanity is that our numbers are growing but the availability of land and fresh water for agriculture is not. This problem is being exacerbated by climate change-induced ...increases in drought, and other abiotic stresses. Stress-resistant crops are needed to ensure yield stability under stress conditions and to minimize the environmental impacts of crop production. Evolution has created thousands of species of naturally stress-resistant plants (NSRPs), some of which have already been subjected to human domestication and are considered minor crops. Broader cultivation of these minor crops will diversify plant agriculture and the human diet, and will therefore help improve global food security and human health. More research should be directed toward understanding and utilizing NSRPs. Technologies are now available that will enable researchers to rapidly improve the genetics of NSRPs, with the goal of increasing NSRP productivity while retaining NSRP stress resistance and nutritional value.
Previous studies on base editing in dicots showed that improvement of nCas9 expression level could significantly increase the editing efficiency (Kang et al., 2018). ...we sought to optimize pCXPE01 ...to improve editing efficiency by increasing nCas9‐MMLV expression level. According to the sequencing results, we detected desired edits at two genes, ALS2 and PDS1. ...no plant phenotype results were reported in these studies. ...our prime‐edited T0 tomato plants were chimeras and did not display any obvious phenotypes (Figure 1e, g and h). ...for both monocots and dicots, assessment of the utility of prime editing awaits future analysis of large populations of edited lines and their off‐springs.
Recently, DNA methylation was proposed to regulate fleshy fruit ripening. Fleshy fruits can be distinguished by their ripening process as climacteric fruits, such as tomatoes, or non-climacteric ...fruits, such as strawberries. Tomatoes undergo a global decrease in DNA methylation during ripening, due to increased expression of a DNA demethylase gene. The dynamics and biological relevance of DNA methylation during the ripening of non-climacteric fruits are unknown.
Here, we generate single-base resolution maps of the DNA methylome in immature and ripe strawberry. We observe an overall loss of DNA methylation during strawberry fruit ripening. Thus, ripening-induced DNA hypomethylation occurs not only in climacteric fruit, but also in non-climacteric fruit. Application of a DNA methylation inhibitor causes an early ripening phenotype, suggesting that DNA hypomethylation is important for strawberry fruit ripening. The mechanisms underlying DNA hypomethylation during the ripening of tomato and strawberry are distinct. Unlike in tomatoes, DNA demethylase genes are not upregulated during the ripening of strawberries. Instead, genes involved in RNA-directed DNA methylation are downregulated during strawberry ripening. Further, ripening-induced DNA hypomethylation is associated with decreased siRNA levels, consistent with reduced RdDM activity. Therefore, we propose that a downregulation of RdDM contributes to DNA hypomethylation during strawberry ripening.
Our findings provide new insight into the DNA methylation dynamics during the ripening of non-climacteric fruit and suggest a novel function of RdDM in regulating an important process in plant development.
DNA methylation is an important epigenetic mark involved in diverse biological processes. In plants, DNA methylation can be established through the RNA-directed DNA methylation pathway, an RNA ...interference pathway for transcriptional gene silencing （TGS）, which requires 24-nt small interfering RNAs. In mammals, de novo DNA methylation occurs primarily at two developmental stages： during early embryogenesis and during gametogenesis. While it is not clear whether establishment of DNA methylation patterns in mammals involves RNA interference in general, de novo DNA methylation and suppression of transposons in germ cells require 24-32-nt piwi-interacting small RNAs. DNA methylation status is dynamically regulated by DNA methylation and demethylation reactions. In plants, active DNA demethylation relies on the repressor of silencing 1 family of bifunctional DNA glycosylases, which remove the 5-methylcytosine base and then cleave the DNA backbone at the abasic site, initiating a base excision repair （BER） pathway. In animals, multiple mechanisms of active DNA demethylation have been proposed, including a deaminase- and DNA glycosylase-initiated BER pathway. New information concerning the effects of various histone modifications on the establishment and maintenance of DNA methylation has broadened our understanding of the regulation of DNA methylation. The function of DNA methylation in plants and animals is also discussed in this review.
The widespread agricultural problem of pre-harvest sprouting (PHS) could potentially be overcome by improving seed dormancy. Here, we report that miR156, an important grain yield regulator, also ...controls seed dormancy in rice. We found that mutations in one MIR156 subfamily enhance seed dormancy and suppress PHS with negligible effects on shoot architecture and grain size, whereas mutations in another MIR156 subfamily modify shoot architecture and increase grain size but have minimal effects on seed dormancy. Mechanistically, mir156 mutations enhance seed dormancy by suppressing the gibberellin (GA) pathway through de-represssion of the miR156 target gene Ideal Plant Architecture 1 (IPA1), which directly regulates multiple genes in the GA pathway. These results provide an effective method to suppress PHS without compromising productivity, and will facilitate breeding elite crop varieties with ideal plant architectures.
PDF
