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  1. Abstract The highly diverse Solanaceae family contains several widely studied models and crop species. Fully exploring, appreciating, and exploiting this diversity requires additional model systems. Particularly promising are orphan fruit crops in the genus Physalis, which occupy a key evolutionary position in the Solanaceae and capture understudied variation in traits such as inflorescence complexity, fruit ripening and metabolites, disease and insect resistance, self-compatibility, and most notable, the striking inflated calyx syndrome (ICS), an evolutionary novelty found across angiosperms where sepals grow exceptionally large to encapsulate fruits in a protective husk. We recently developed transformation and genome editing in Physalis grisea (groundcherry). However, to systematically explore and unlock the potential of this and related Physalis as genetic systems, high-quality genome assemblies are needed. Here, we present chromosome-scale references for P. grisea and its close relative Physalis pruinosa and use these resources to study natural and engineered variations in floral traits. We first rapidly identified a natural structural variant in a bHLH gene that causes petal color variation. Further, and against expectations, we found that CRISPR–Cas9-targeted mutagenesis of 11 MADS-box genes, including purported essential regulators of ICS, had no effect on inflation. In a forward genetics screen, we identified huskless, which lacksmore »ICS due to mutation of an AP2-like gene that causes sepals and petals to merge into a single whorl of mixed identity. These resources and findings elevate Physalis to a new Solanaceae model system and establish a paradigm in the search for factors driving ICS.« less
    Free, publicly-accessible full text available October 21, 2023
  2. Abstract

    Poleroviruses, enamoviruses, and luteoviruses are icosahedral, positive sense RNA viruses that cause economically important diseases in food and fiber crops. They are transmitted by phloem-feeding aphids in a circulative manner that involves the movement across and within insect tissues. The N-terminal portion of the viral readthrough domain (NRTD) has been implicated as a key determinant of aphid transmission in each of these genera. Here, we report crystal structures of theNRTDs from the poleroviruses turnip yellow virus (TuYV) and potato leafroll virus (PLRV) at 1.53-Å and 2.22-Å resolution, respectively. These adopt a two-domain arrangement with a unique interdigitated topology and form highly conserved dimers that are stabilized by a C-terminal peptide that is critical for proper folding. We demonstrate that the PLRVNRTD can act as an inhibitor of virus transmission and identifyNRTD mutant variants that are lethal to aphids. Sequence conservation argues that enamovirus and luteovirusNRTDs will follow the same structural blueprint, which affords a biological approach to block the spread of these agricultural pathogens in a generalizable manner.

  3. Free, publicly-accessible full text available March 1, 2023
  4. Free, publicly-accessible full text available April 1, 2023
  5. Most plant roots have multiple cortex layers that make up the bulk of the organ and play key roles in physiology, such as flood tolerance and symbiosis. However, little is known about the formation of cortical layers outside of the highly reduced anatomy of Arabidopsis . Here, we used single-cell RNA sequencing to rapidly generate a cell-resolution map of the maize root, revealing an alternative configuration of the tissue formative transcription factor SHORT-ROOT (SHR) adjacent to an expanded cortex. We show that maize SHR protein is hypermobile, moving at least eight cell layers into the cortex. Higher-order SHR mutants in both maize and Setaria have reduced numbers of cortical layers, showing that the SHR pathway controls expansion of cortical tissue to elaborate anatomical complexity.
  6. Abstract Cytosine base editors (CBEs) are promising tools for precise genome editing in plants. It is important to investigate potential off-target effects of an efficient CBE at the genome and transcriptome levels in a major crop. Based on comparison of five cytidine deaminases and two different promoters for expressing sgRNAs, we tested a highly efficient A3A/Y130F-BE3 system for efficient C-to-T base editing in tomato (Solanum lycopersicum). We then conducted whole-genome sequencing (WGS) of four base-edited tomato plants, three GFP-expressing control plants, and two wild-type (WT) plants. The sequencing depths ranged from 25X to 49X with read mapping rates above 97%. No sgRNA-dependent off-target mutations were detected. Our data show an average of ∼1000 single nucleotide variations (SNVs) and ∼100 insertions and deletions (indels) per GFP control plant. Base-edited plants had on average elevated levels of SNVs (∼1250) and indels (∼300) per plant. On average, about 200 more C-to-T (G-to-A) mutations were found in a base-edited plant than a GFP control plant, suggesting some level of sgRNA-independent off-target effects, though the difference is not statistically significant. We also conducted RNA sequencing (RNA-seq) of the same four base-edited plants and three GFP control plants. An average of ∼200 RNA SNVs was discoveredmore »per plant for either base-edited or GFP control plants. Furthermore, no specific enrichment of C-to-U mutations can be found in the base-edited plants. Hence, we cannot find any evidence for bona fide off-target mutations by A3A/Y130F-BE3 at the transcriptome level.« less
  7. Fruit softening is a key component of the irreversible ripening program, contributing to the palatability necessary for frugivore-mediated seed dispersal. The underlying textural changes are complex and result from cell wall remodeling and changes in both cell adhesion and turgor. While a number of transcription factors (TFs) that regulate ripening have been identified, these affect most canonical ripening-related physiological processes. Here, we show that a tomato fruit ripening–specific LATERAL ORGAN BOUNDRIES ( LOB ) TF, SlLOB1 , up-regulates a suite of cell wall–associated genes during late maturation and ripening of locule and pericarp tissues. SlLOB1 repression in transgenic fruit impedes softening, while overexpression throughout the plant under the direction of the 35s promoter confers precocious induction of cell wall gene expression and premature softening. Transcript and protein levels of the wall-loosening protein EXPANSIN1 ( EXP1 ) are strongly suppressed in Sl LOB1 RNA interference lines, while EXP1 is induced in Sl LOB1 -overexpressing transgenic leaves and fruit. In contrast to the role of ethylene and previously characterized ripening TFs, which are comprehensive facilitators of ripening phenomena including softening, Sl LOB1 participates in a regulatory subcircuit predominant to cell wall dynamics and softening.
  8. null (Ed.)