Tomato and mini-cucumber tolerance to photoperiodic injury involves photorespiration and the engagement of nighttime cyclic electron flow from dynamic LEDs.

Controlled environment agriculture (CEA) is critical for achieving year-round food security in many regions of the world. CEA is a resource-intensive endeavor, with lighting consuming a large fraction of the energy. To lessen the burden on the grid and save costs, an extended photoperiod strategy can take advantage of off-peak time-of-day options from utility suppliers. However, extending the photoperiod limits crop production morphologically and physiologically if pushed too long. Here, we present a continuous-light dynamic light-emitting diode (LED) strategy (involving changes in spectra, intensity, and timing), that overcomes these limitations. We focused on tomato, a well described photoperiodic injury-sensitive species, and mini-cucumber, a photoperiodic injury-tolerant species to first assess morphological responses under control (16-h photoperiod, unchanging spectrum), constant (24-h photoperiod, unchanging spectrum), and two variations of a dynamic LED strategy, dynamic 1 (16-h "day", 3-h "peak", 8-h "night" spectra) and dynamic 2 (20-h "day", 5-h "peak", 4-h "night" spectra). Next, we tested the hypothesis of photorespiration's involvement in photoperiodic injury by using a leaf gas exchange coupled with chlorophyll fluorescence protocol. We further explored Adenosine triphosphate (ATP): Nicotinamide adenine dinucleotide phosphate (NADPH) ratio supply/demand responses by probing photosynthetic electron flow and proton flow with the MultispeQ instrument. We found canopy architecture can be tuned by minor variations of the same dynamic LED strategy, and we highlight dynamic 1 as the optimal choice for both tomato and mini-cucumber as it improved biomass/architecture and first-yield, respectively. A central discovery was that dynamic 1 had a significantly higher level of photorespiration than control, for both species. Unexpectedly, photorespiration was comparable between species under the same treatments, except under constant. However, preliminary data on a fully tolerant tomato genotype grown under constant treatment upregulated photorespiration similar to mini-cucumber. These results suggest that photoperiodic injury tolerance involves a sustained higher level of photorespiration under extended photoperiods. Interestingly, diurnal MultispeQ measurements point to the importance of cyclic electron flow at subjective nighttime that may also partially explain why dynamic LED strategies mitigate photoperiodic injury. We propose an ontology of photoperiodic injury involving photorespiration, triose phosphate utilization, peroxisomal H2O2-catalase balance, and a circadian external coincidence model of sensitivity that initiates programmed cell death.

Tissue culture coupled with a gas exchange system offers new perspectives on phenotyping the developmental biology of Solanum lycopersicum L. cv. 'MicroTom'.

Solanum lycopersicum L. cv. 'Microtom' (MicroTom) is a model organism with a relatively rapid life cycle, and wide library of genetic mutants available to study different aspects of plant development. Despite its small stature, conventional MicroTom research often requires expensive growth cabinets and/or expansive greenhouse space, limiting the number of experimental and control replications needed for experiments, and can render plants susceptible to pests and disease. Thus, alternative experimental approaches must be devised to reduce the footprint of experimental units and limit the occurrence problematic confounding variables. Here, tissue culture is presented as a powerful option for MicroTom research that can quell the complications associated with conventional MicroTom research methods. A previously established, non-invasive, analytical tissue culture system is used to compare in vitro and conventionally produced MicroTom by assessing photosynthesis, respiration, diurnal carbon gain, and fruit pigments. To our knowledge, this is the first publication that measures in vitro MicroTom fruit pigments and compares diurnal photosynthetic/respiration responses to abiotic factors between in vitro and ex vitro MicroTom. Comparable trends would validate tissue culture as a new benchmark method in MicroTom research, as it is like Arabidopsis, allowing replicable, statistically valid, high throughput genotyping and selective phenotyping experiments. Combining the model plant MicroTom with advanced tissue culture methods makes it possible to study bonsai-style MicroTom responses to light, temperature, and atmospheric stimuli in the absence of confounding abiotic stress factors that would otherwise be unachievable using conventional methods.

Novel mutations and electrophysiologic findings in RGS9- and R9AP-associated retinal dysfunction (Bradyopsia).

PURPOSE: To examine the phenotypes of 8 patients with evidence of cone dysfunction and normal color vision (characteristic features of both oligocone trichromacy and bradyopsia), and subsequently to screen RGS9 and R9AP for disease-causing mutations. DESIGN: Retrospective case series. PARTICIPANTS: Eight affected individuals from 7 families. METHODS: Ophthalmologic examination, color vision testing, fundus photography, and detailed electrophysiologic assessment were undertaken. Blood samples were taken for DNA extraction from affected subjects and, where possible, unaffected relatives. Mutation screening of RGS9 and R9AP was performed. MAIN OUTCOME MEASURES: Detailed clinical, electrophysiologic, and molecular genetic findings. RESULTS: All 8 patients had normal ocular examination results, with visual acuity ranging from 6/12 to 6/18. Four subjects were found to harbor mutations in RGS9 or R9AP, with 3 of the identified sequence variants being novel. Three subjects, 2 Pakistani sisters and an Afghani female, had mutations in R9AP. A novel homozygous nonsense mutation, p.G205fs, was identified in the simplex case, and a second novel homozygous in-frame deletion, p.D32_Q34del, was found in the 2 sisters. The remaining patient, a British male, had a compound heterozygous mutation in RGS9 (p.R128X/p.W299R). The mutation p.R128X represents the first nonsense mutation reported in RGS9. The 4 mutation-positive subjects had concordant characteristic previously described electrophysiologic findings that were not present in the 4 individuals in whom mutations were not identified. Novel findings associated with these mutation-positive patients included that they all showed electroretinogram evidence of severe cone system dysfunction under photopic conditions but normal cone function to a red flash under scotopic conditions. Such findings seem unique for the disorder. CONCLUSIONS: This is the first report describing a nonsense mutation in RGS9. We have established novel electrophysiologic observations associated with RGS9 and R9AP mutations, including those relating to dark-adapted cone function and S-cone function. Patients with either RGS9/R9AP mutations (bradyopsia) or oligocone trichromacy have very similar clinical phenotypes, characterized by stationary cone dysfunction, mild photophobia, normal color vision, lack of nystagmus, and normal fundi. The distinctive electrophysiologic features associated with RGS9 and R9AP mutations enable directed genetic screening. FINANCIAL DISCLOSURE(S): The author(s) have no proprietary or commercial interest in any materials discussed in this article.

The loss of the PDE6 deactivating enzyme, RGS9, results in precocious light adaptation at low light levels.

The GTPase activating protein, RGS9-1, is vital for the deactivation and regulation of the phototransduction cascade (C. K. Chen et al., 2000; C. W. Cowan, R. N. Fariss, I. Sokal, K. Palczewski, & T. G. Wensel, 1998; W. He, C. W. Cowan, & T. G. Wensel, 1998; A. L. Lyubarsky et al., 2001). Its loss through genetic defects in humans has been linked to a slow recovery to changes in illumination (K. M. Nishiguchi et al., 2004). Such a deficit is to be expected because RGS9-1 normally speeds up the deactivation of the activated phosphodiesterase effector molecule, PDE6*, and thus accelerates the turning off of the visual response. Paradoxically, however, we find that the cone response in an observer lacking RGS9-1 is faster at lower light levels than it is in a normal observer. Though surprising, this result is nonetheless consistent with molecular models of light adaptation (e.g., E. N. Pugh, S. Nikonov, & T. D. Lamb, 1999), which predict that the excess of PDE6* resulting from the loss of RGS9-1 will shorten the visual integration time and speed up the visual response at inappropriately low light levels. The gain in speed caused by the superfluity of PDE6* at lower light levels compensates for the loss caused by its slow deactivation; thus quickening the response relative to that in the normal. As the light level is increased and the PDE6* concentration in the normal rises relative to that in the observer lacking RGS9-1, the temporal advantage of the latter is soon lost, leaving only the deficit due to delayed deactivation.

Recombination hotspots and block structure of linkage disequilibrium in the human genome exemplified by detailed analysis of PGM1 on 1p31.

The distribution of linkage disequilibrium (LD) in the human genome has important consequences for the design of experiments that infer susceptibility genes for complex disease using association studies. Recent studies have shown a non-random distribution of human meiotic recombination associated with intervening tracts of LD. Little is known about the processes, patterns and frequency of reciprocal meiotic recombination in humans. However, this phenomenon can be better understood by the fine structure analysis of several genomic regions by mapping hotspots and characterizing regions with variable LD. Here, we report clustered hotspot activity with intervening blocks of LD within the human PGM1 gene (1p31) using data derived from meiotic and population studies. Earlier work has suggested a high recombination rate in two regions within the PGM1 gene, site A (exons 4-8) and site B (exons 1A-4). Sequencing of eight individuals across 6 kb of targeted regions in site B identified 18 informative SNPs. Individuals from three distinct populations, Caucasian (n=264), Chinese (n=222) and Vietnamese (n=187), were genotyped, and haplotypes were determined using estimate of haplotypes, ldmax and Arlequin. Allelic association and haplotype analysis in these samples revealed variable recombination rates across PGM1, demonstrating the presence of: (i) three hotspots and (ii) three haplotype blocks. The spatial arrangement of haplotype blocks was identical in all populations studied. The pattern of association within PGM1 represents a region decomposed into small blocks of LD, where increased recombination activity has disrupted the ancestral chromosome. Additionally, crossovers in phased data mapped preferentially to regions where LD collapses, which also overlap with sequence motifs.