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1. t‐SNAREs bind the Rhg1 α‐SNAP and mediate soybean cyst nematode resistance

   https://doi.org/10.1111/tpj.14923  

   Dong, Jia ; Zielinski, Raymond E. ; Hudson, Matthew E. ( August 2020 , The Plant Journal)

   Soybean cyst nematode (SCN;Heterodera glycines) is the largest pathogenic cause of soybean yield loss. TheRhg1locus is the most used and best characterized SCN resistance locus, and contains three genes including one encoding an α‐SNAP protein. Although theRhg1α‐SNAP is known to play an important role in vesicle trafficking and SCN resistance, the protein’s binding partners and the molecular mechanisms underpinning SCN resistance remain unclear. In this report, we show that theRhg1α‐SNAP strongly interacts with two syntaxins of the t‐SNARE family (Glyma.12G194800 and Glyma.16G154200) in yeast and plants; importantly, the genes encoding these syntaxins co‐localize with SCN resistance quantitative trait loci. Fluorescent visualization revealed that the α‐SNAP and the two interacting syntaxins localize to the plasma membrane and perinuclear space in both tobacco epidermal and soybean root cells. The two syntaxins and their two homeologs were mutated, individually and in combination, using the CRISPR‐Cas9 system in the SCN‐resistant Peking and SCN‐susceptible Essex soybean lines. Peking roots with deletions introduced into syntaxin genes exhibited significantly reduced resistance to SCN, confirming that t‐SNAREs are critical to resisting SCN infection. The results presented here uncover a key step in the molecular mechanism of SCN resistance, and will be invaluable to soybean breeders aiming to develop highly SCN‐resistant soybean varieties.

    

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2. The rhg1‐a ( Rhg1 low‐copy) nematode resistance source harbors a copia‐family retrotransposon within the Rhg1‐ encoded α‐SNAP gene

   https://doi.org/10.1002/pld3.164  

   Bayless, Adam M. ; Zapotocny, Ryan W. ; Han, Shaojie ; Grunwald, Derrick J. ; Amundson, Kaela K. ; Bent, Andrew F. ( August 2019 , Plant Direct)

   Soybean growers widely use theResistance toHeteroderaglycines1 (Rhg1) locus to reduce yield losses caused by soybean cyst nematode (SCN).Rhg1is a tandemly repeated four gene block. Two classes of SCN resistance‐conferringRhg1haplotypes are recognized:rhg1‐a(“Peking‐type,” low‐copy number, three or fewerRhg1repeats) andrhg1‐b(“PI 88788‐type,” high‐copy number, four or moreRhg1repeats). Therhg1‐aandrhg1‐bhaplotypes encode α‐SNAP (alpha‐SolubleNSFAttachmentProtein) variants α‐SNAP_Rhg1LC and α‐SNAP_Rhg1HC, respectively, with differing atypical C‐terminal domains, that contribute to SCN resistance. Here we report thatrhg1‐asoybean accessions harbor a copia retrotransposon within theirRhg1 Glyma.18G022500(α‐SNAP‐encoding) gene. We termed this retrotransposon “RAC,” forRhg1alpha‐SNAPcopia. Soybean carries multipleRAC‐like retrotransposon sequences. TheRhg1 RACinsertion is in theGlyma.18G022500genes of all truerhg1‐ahaplotypes we tested and was not detected in any examinedrhg1‐borRhg1_WT(single‐copy) soybeans.RACis an intact element residing within intron 1, anti‐sense to therhg1‐a α‐SNAPopen reading frame.RAChas intrinsic promoter activities, but overt impacts ofRACon transgenic α‐SNAP_Rhg1LC mRNA and protein abundance were not detected. From the nativerhg1‐a RAC⁺genomic context, elevated α‐SNAP_Rhg1LC protein abundance was observed in syncytium cells, as was previously observed for α‐SNAP_Rhg1HC (whoserhg1‐bdoes not carryRAC). Using a SoySNP50K SNP corresponding withRACpresence, just ~42% of USDA accessions bearing previously identifiedrhg1‐aSoySNP50K SNP signatures harbor theRACinsertion. Subsequent analysis of several of these putativerhg1‐aaccessions lackingRACrevealed that none encodedα‐SNAP_Rhg1LC, and thus, they are notrhg1‐a.rhg1‐ahaplotypes are of rising interest, withRhg4, for combating SCN populations that exhibit increased virulence against the widely usedrhg1‐bresistance. The present study reveals another unexpected structural feature of manyRhg1loci, and a selectable feature that is predictive ofrhg1‐ahaplotypes.

    

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3. Structural and functional analysis of two SHMT8 variants associated with soybean cyst nematode resistance

   https://doi.org/10.1111/febs.16971  

   Korasick, David A. ; Owuocha, Luckio F. ; Kandoth, Pramod K. ; Tanner, John J. ; Mitchum, Melissa G. ; Beamer, Lesa J. ( October 2023 , The FEBS Journal)

   Two amino acid variants in soybean serine hydroxymethyltransferase 8 (SHMT8) are associated with resistance to the soybean cyst nematode (SCN), a devastating agricultural pathogen with worldwide economic impacts on soybean production. SHMT8 is a cytoplasmic enzyme that catalyzes the pyridoxal 5‐phosphate‐dependent conversion of serine and tetrahydrofolate (THF) to glycine and 5,10‐methylenetetrahydrofolate. A previous study of the P130R/N358Y double variant of SHMT8, identified in the SCN‐resistant soybean cultivar (cv.) Forrest, showed profound impairment of folate binding affinity and reduced THF‐dependent enzyme activity, relative to the highly active SHMT8 in cv. Essex, which is susceptible to SCN. Given the importance of SCN‐resistance in soybean agriculture, we report here the biochemical and structural characterization of the P130R and N358Y single variants to elucidate their individual effects on soybean SHMT8. We find that both single variants have reduced THF‐dependent catalytic activity relative to Essex SHMT8 (10‐ to 50‐fold decrease ink_cat/K_m) but are significantly more active than the P130R/N368Y double variant. The kinetic data also show that the single variants lack THF‐substrate inhibition as found in Essex SHMT8, an observation with implications for regulation of the folate cycle. Five crystal structures of the P130R and N358Y variants in complex with various ligands (resolutions from 1.49 to 2.30 Å) reveal distinct structural impacts of the mutations and provide new insights into allosterism. Our results support the notion that the P130R/N358Y double variant in Forrest SHMT8 produces unique and unexpected effects on the enzyme, which cannot be easily predicted from the behavior of the individual variants.

    

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4. Movement and Motion of Soybean Cyst Nematode Heterodera glycines Populations and Individuals in Response to Abamectin

   https://doi.org/10.1094/PHYTO-10-17-0339-R  

   Jensen, Jared P. ; Beeman, Augustine Q. ; Njus, Zach L. ; Kalwa, Upender ; Pandey, Santosh ; Tylka, Gregory L. ( July 2018 , Phytopathology®)

   Two new in vitro methods were developed to analyze plant-parasitic nematode behavior, at the population and the individual organism levels, through time-lapse image analysis. The first method employed a high-resolution flatbed scanner to monitor the movement of a population of nematodes over a 24-h period at 25°C. The second method tracked multiple motion parameters of individual nematodes on a microscopic scale, using a high-speed camera. Changes in movement and motion of second-stage juveniles (J2) of the soybean cyst nematode Heterodera glycines Ichinohe were measured after exposure to a serial dilution of abamectin (0.1 to 100 μg/ml). Movement and motion of H. glycines were significantly reduced as the concentration of abamectin increased. The effective range of abamectin to inhibit movement and motion of H. glycines J2 was between 1.0 and 10 μg/ml. Proof-of-concept experiments for both methods produced one of the first in vitro sensitivity studies of H. glycines to abamectin. The two methods developed allow for higher-throughput analysis of nematode movement and motion and provide objective and data-rich measurements that are difficult to achieve from conventional microscopic laboratory methods. 

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5. Growing Degree-Day Measurement of Cyst Germination Rates in the Toxic Dinoflagellate Alexandrium catenella

   https://doi.org/10.1128/aem.02518-21  

   Fischer, Alexis D. ; Brosnahan, Michael L. ( June 2022 , Applied and Environmental Microbiology)

   Rudi, Knut (Ed.)

   ABSTRACT Blooms of many dinoflagellates, including several harmful algal bloom (HAB) species, are seeded and revived through the germination of benthic resting cysts. Temperature is a key determinant of cysts’ germination rate, and temperature–germination rate relationships are therefore fundamental to understanding species’ germling cell production, cyst bed persistence, and resilience to climate warming. This study measured germination by cysts of the HAB dinoflagellate Alexandrium catenella using a growing degree-day ( DD ) approach that accounts for the time and intensity of warming above a critical temperature. Time courses of germination at different temperatures were fit to lognormal cumulative distribution functions for the estimation of the median days to germination. As temperature increased, germination times decreased hyperbolically. DD scaling collapsed variability in germination times between temperatures after cysts were oxygenated. A parallel experiment demonstrated stable temperature–rate relationships in cysts collected during different phases of seasonal temperature cycles in situ over three years. DD scaling of the results from prior A. catenella germination studies showed consistent differences between populations across a wide range of temperatures and suggests selective pressure for different germination rates. The DD model provides an elegant approach to quantify and compare the temperature dependency of germination among populations, between species, and in response to changing environmental conditions. IMPORTANCE Germination by benthic life history stages is the first step of bloom initiation in many, diverse phytoplankton species. This study outlines a growing degree-day ( DD ) approach for comparing the temperature dependence of germination rates measured in different populations. Germination by cysts of Alexandrium catenella , a harmful algal bloom dinoflagellate that causes paralytic shellfish poisoning, is shown to require a defined amount of warming, measured in DD after cysts are aerated. Scaling by DD , the time integral of temperature difference from a critical threshold, enabled direct comparison of rates measured at different temperatures and in different studies. 

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