Shapes of edible plant organs vary dramatically among and within crop plants. To explain and ultimately employ this variation towards crop improvement, we determined the genetic, molecular and cellular bases of fruit shape diversity in tomato. Through positional cloning, protein interaction studies, and genome editing, we report that OVATE Family Proteins and TONNEAU1 Recruiting Motif proteins regulate cell division patterns in ovary development to alter final fruit shape. The physical interactions between the members of these two families are necessary for dynamic relocalization of the protein complexes to different cellular compartments when expressed in tobacco leaf cells. Together with data from other domesticated crops and model plant species, the protein interaction studies provide possible mechanistic insights into the regulation of morphological variation in plants and a framework that may apply to organ growth in all plant species.

Domestication of fruit and vegetables resulted in a huge diversity of shapes and sizes of the produce. Selections that took place over thousands of years of alleles that increased fruit weight and altered shape for specific culinary uses provide a wealth of resources to study the molecular bases of this diversity. Tomato (Solanum lycopersicum) evolved from a wild ancestor (S. pimpinellifolium) bearing small and round edible fruit. Molecular genetic studies led to the identification of two genes selected for fruit weight: FW2.2 encoding a member of the Cell Number Regulator family; and FW3.2 encoding a P450 enzyme and the ortholog of KLUH. Four genes were identified that were selected for fruit shape: SUN encoding a member of the IQD family of calmodulin-binding proteins leading to fruit elongation; OVATE encoding a member of the OVATE family proteins involved in transcriptional repression leading to fruit elongation; LC encoding most likely the ortholog of WUSCHEL controlling meristem size and locule number; FAS encoding a member in the YABBY family controlling locule number leading to flat or oxheart shape. For this article, we will provide an overview of the putative function of the known genes, when during floral and fruit development they are hypothesized to act and their potential importance in regulating morphological diversity in other fruit and vegetable crops.

Accumulation of capsaicinoids in the placental tissue of ripening chile (Capsicum spp.) fruit follows the coordinated expression of multiple biosynthetic enzymes producing the substrates for capsaicin synthase. Transcription factors are likely agents to regulate expression of these biosynthetic genes. Placental RNAs from habanero fruit (C. chinense) were screened for expression of candidate transcription factors; with two candidate genes identified, both in the ERF family of transcription factors. Characterization of these transcription factors, Erf and Jerf, in nine chile cultivars with distinct capsaicinoid contents demonstrated a correlation of expression with pungency. Amino acid variants were observed in both ERF and JERF from different chile cultivars; none of these changes involved the DNA binding domains. Little to no transcription of Erf was detected in non-pungent C. annuum or C. chinense mutants. This correlation was characterized at an individual fruit level in a set of jalapeño (C. annuum) lines again with distinct and variable capsaicinoid contents. Both Erf and Jerf are expressed early in fruit development, 16–20 days post-anthesis, at times prior to the accumulation of capsaicinoids in the placental tissues. These data support the hypothesis that these two members of the complex ERF family participate in regulation of the pungency phenotype in chile.

The spindle assembly checkpoint (SAC) delays mitotic progression until all sister chromatid pairs achieve bi-orientation, and while the SAC can maintain mitotic arrest for extended periods, moderate delays in mitotic progression have significant effects on the resulting daughter cells. Here we show that when retinal-pigmented epithelial (RPE1) cells experience mitotic delay, there is a time-dependent increase in centrosome fragmentation and centriole disengagement. While most cells with disengaged centrioles maintain spindle bipolarity, clustering of disengaged centrioles requires the kinesin-14, HSET. Centrosome fragmentation and precocious centriole disengagement depend on separase and anaphase-promoting complex/cyclosome (APC/C) activity, which also triggers the acquisition of distal appendage markers on daughter centrioles and the loss of procentriolar markers. Together, these results suggest that moderate delays in mitotic progression trigger the initiation of centriole licensing through centriole disengagement, at which point the ability to maintain spindle bipolarity becomes a function of HSET-mediated spindle pole clustering.

Aberrant regulation of metabolic kinases by altered redox homeostasis substantially contributes to aging and various diseases, such as diabetes. We found that the catalytic activity of a conserved family of fructosamine-3-kinases (FN3Ks), which are evolutionarily related to eukaryotic protein kinases, is regulated by redox-sensitive cysteine residues in the kinase domain. The crystal structure of the FN3K homolog from Arabidopsis thaliana revealed that it forms an unexpected strand-exchange dimer in which the ATP-binding P-loop and adjoining β strands are swapped between two chains in the dimer. This dimeric configuration is characterized by strained interchain disulfide bonds that stabilize the P-loop in an extended conformation. Mutational analysis and solution studies confirmed that the strained disulfides function as redox “switches” to reversibly regulate the activity and dimerization of FN3K. Human FN3K, which contains an equivalent P-loop Cys, was also redox sensitive, whereas ancestral bacterial FN3K homologs, which lack a P-loop Cys, were not. Furthermore, CRISPR-mediated knockout of FN3K in human liver cancer cells altered the abundance of redox metabolites, including an increase in glutathione. We propose that redox regulation evolved in FN3K homologs in response to changing cellular redox conditions. Our findings provide insights into the origin and evolution of redox regulation in the protein kinase superfamily and may open new avenues for targeting human FN3K in diabetic complications.

The carotenoid content of fresh fruits, like chiles or peppers (Capsicum annuum L.), is a desirable fruit quality trait because these compounds increase the nutritional value of the fruit. Carotenoids in general serve as antioxidants, whereas specific carotenoids are pro-vitamin A types and yet others are necessary for retinal pigments. In the plant, carotenoids function to harvest light energy during photosynthesis, act as antioxidants in multiple cell types, and pigment fruit and flowers to attract pollinators and seed dispersal agents. All of these cells presumably accumulate carotenoids through the same biosynthetic pathway. We investigated the relationship between light levels in the growth environment and the carotenoid levels that accumulated in mature fruit and leaves. Three chile cultivars with orange fruit, ‘Fogo’, ‘Orange Grande’, and ‘NuMex Sunset’, were grown under three different light conditions, shaded greenhouse, unshaded greenhouse, and field in Las Cruces, NM. Foliar carotenoid increased approximately twofold with increased light, whereas carotenoid content in fruit decreased two- to threefold with increased light. All cultivars showed identical trends with light despite having cultivar-specific carotenoid accumulation patterns in their fruit.

words)Aberrant regulation of metabolic kinases by altered redox homeostasis is a major contributing factor in aging and disease such as diabetes. However, the biochemical mechanisms by which metabolic kinases are regulated under oxidative stress is poorly understood. In this study, we demonstrate that the catalytic activity of a conserved family of Fructosamine-3-kinases (FN3Ks), which are evolutionarily related to eukaryotic protein kinases (ePKs), are regulated by redox active cysteines in the kinase domain. By solving the crystal structure of FN3K homolog from Arabidopsis thaliana (AtFN3K), we demonstrate that it forms an unexpected strandexchange dimer in which the ATP binding P-loop and adjoining beta strands are swapped between two chains in the dimer. This dimeric configuration is characterized by strained inter-chain disulfide bonds that stabilize the P-loop in an extended conformation and tethers the substratebinding lobes in a unique inactive conformation. Mutational analysis and solution studies confirm that the strained disulfides function as redox "switches" to reversibly regulate FN3K activity and dimerization. Consistently, we find that human FN3K, which contains an equivalent P-loop Cys, is also redox sensitive, whereas ancestral bacterial FN3K homologs, which lack a P-loop Cys, are not. Furthermore, CRISPR knockout of FN3K in human HepG2 cells results in significant upregulation of redox metabolites including glutathione. We propose that redox regulation evolved progressively in FN3Ks in response to changing cellular redox conditions. Our studies provide important new insights into the origin and evolution of redox regulation in the protein kinase superfamily and open new avenues for targeting human FN3K in diabetic complications.

Summary The mechanisms that regulate the vast diversity of plant organ shapes such as the fruit remain to be fully elucidated. TONNEAU1 Recruiting Motif proteins (TRMs) have been implicated in the control of organ shapes in a number of plant species, including tomato. However, the role of many of them is unknown. TRMs interact with Ovate Family Proteins (OFPs) via the M8 domain. However, the in planta function of the TRM‐OFP interaction in regulating shape is unknown. We used CRISPR/Cas9 to generate knockout mutants in TRM proteins from different subclades and in‐frame mutants within the M8 domain to investigate their roles in organ shape and interactions with OFPs. Our findings indicate that TRMs impact organ shape along both the mediolateral and proximo‐distal axes of growth. Mutations in Sltrm3/4 and Sltrm5 act additively to rescue the elongated fruit phenotype of ovate/Slofp20 (o/s) to a round shape. Contrary, mutations in Sltrm19 and Sltrm17/20a result in fruit elongation and further enhance the obovoid phenotype in the o/s mutant. This study supports a combinatorial role of the TRM‐OFP regulon where OFPs and TRMs expressed throughout development have both redundant and opposing roles in regulating organ shape.