The loss of function of HEL, which encodes a cellulose synthase interactive protein, causes helical and vine-like growth of tomato

Helical growth is an economical way for plant to obtain resources. The classic microtubule–microfibril alignment model of Arabidopsis helical growth involves restriction of the appropriate orientation of cellulose microfibrils appropriately in the cell walls. However, the molecular mechanism underlying tomato helical growth remains unknown. Here, we identified a spontaneous tomato helical (hel) mutant with right-handed helical cotyledons and petals but left-handed helical stems and true leaves. Genetic analysis revealed that the hel phenotype was controlled by a single recessive gene. Using map-based cloning, we cloned the HEL gene, which encodes a cellulose interacting protein homologous to CSI1 of Arabidopsis. We identified a 27 bp fragment replacement that generated a premature stop codon. Transgenic experiments showed that the helical growth phenotype could be restored by the allele of this gene from wild-type Pyriforme. In contrast, the knockout mutation of HEL in Pyriforme via CRISPR/Cas9 resulted in helical growth. These findings shed light on the molecular control of the helical growth of tomato.

A moonlighting role for enzymes of glycolysis in the co-localization of mitochondria and chloroplasts

Glycolysis is one of the primordial pathways of metabolism, playing a pivotal role in energy metabolism and biosynthesis. Glycolytic enzymes are known to form transient multi-enzyme assemblies. Here we examine the wider protein-protein interactions of plant glycolytic enzymes and reveal a moonlighting role for specific glycolytic enzymes in mediating the co-localization of mitochondria and chloroplasts. Knockout mutation of phosphoglycerate mutase or enolase resulted in a significantly reduced association of the two organelles. We provide evidence that phosphoglycerate mutase and enolase form a substrate-channelling metabolon which is part of a larger complex of proteins including pyruvate kinase. These results alongside a range of genetic complementation experiments are discussed in the context of our current understanding of chloroplast-mitochondrial interactions within photosynthetic eukaryotes.

Programmed knockout mutation of liver fluke granulin attenuates virulence of infection-induced hepatobiliary morbidity

Infection with the food-borne liver fluke Opisthorchis viverrini is the principal risk factor (IARC Working Group on the Evaluation of Carcinogenic Risks to Humans, 2012) for cholangiocarcinoma (CCA) in the Lower Mekong River Basin countries including Thailand, Lao PDR, Vietnam and Cambodia. We exploited this link to explore the role of the secreted growth factor termed liver fluke granulin (Ov-GRN-1) in pre-malignant lesions by undertaking programmed CRISPR/Cas9 knockout of the Ov-GRN-1 gene from the liver fluke genome. Deep sequencing of amplicon libraries from genomic DNA of gene-edited parasites revealed Cas9-catalyzed mutations within Ov-GRN-1. Gene editing resulted in rapid depletion of Ov-GRN-1 transcripts and the encoded Ov-GRN-1 protein. Gene-edited parasites colonized the biliary tract of hamsters and developed into adult flukes, but the infection resulted in reduced pathology as evidenced by attenuated biliary hyperplasia and fibrosis. Not only does this report pioneer programmed gene-editing in parasitic flatworms, but also the striking, clinically-relevant pathophysiological phenotype confirms the role for Ov-GRN-1 in virulence morbidity during opisthorchiasis.

Ontogenetic Changes in Auxin Biosynthesis and Distribution Determine the Organogenic Activity of the Shoot Apical Meristem in pin1 Mutants

In the shoot apical meristem (SAM) of Arabidopsis, PIN1-dependent polar auxin transport (PAT) regulates two crucial developmental processes: organogenesis and vascular system formation. However, the knockout mutation in the PIN1 gene does not fully inhibit these two processes. Therefore, we investigated a potential source of auxin for organogenesis and vascularization during inflorescence stem development. We analyzed auxin distribution in wild-type (WT) and pin1 mutant plants using a refined protocol of auxin immunolocalization; auxin activity, with the response reporter pDR5:GFP; and expression of auxin biosynthesis genes YUC1 and YUC4. Our results revealed that regardless of the functionality of PIN1-mediated PAT, auxin is present in the SAM and vascular strands. In WT plants, auxin always accumulates in all cells of the SAM, whereas in pin1 mutants, its localization within the SAM changes ontogenetically and is related to changes in the structure of the vascular system, organogenic activity of SAM, and expression levels of YUC1 and YUC4 genes. Our findings indicate that the presence of auxin in the meristem of pin1 mutants is an outcome of at least two PIN1-independent mechanisms: acropetal auxin transport from differentiated tissues with the use of vascular strands and auxin biosynthesis within the SAM.