Rice (Oryza sativa) TIR1 and 5′adamantyl-IAA Significantly Improve the Auxin-Inducible Degron System in Schizosaccharomyces pombe

The auxin-inducible degron (AID) system is a powerful tool to induce targeted degradation of proteins in eukaryotic model organisms. The efficiency of the existing Schizosaccharomyces pombe AID system is limited due to the fusion of the F-box protein TIR1 protein to the SCF component, Skp1 (Skp1-TIR1). Here, we report an improved AID system for S. pombe that uses the TIR1 from Oryza sativa (OsTIR1) not fused to Skp1. Furthermore, we demonstrate that degradation efficiency can be improved by pairing an OsTIR1 auxin-binding site mutant, OsTIR1F74A, with an auxin analogue, 5′adamantyl-IAA (AID2). We provide evidence for the enhanced functionality of the OsTIR1 AID and AID2 systems by application to the essential DNA replication factor Mcm4 and to a non-essential recombination protein, Rad52. Unlike AID, no detectable auxin-independent depletion of AID-tagged proteins was observed using AID2.

Identification and Expression Analysis of Gretchen Hagen 3 (GH3) in Kiwifruit (Actinidia chinensis) During Postharvest Process

In plants, the Gretchen GH3 (GH3) protein is involved in free auxin (IAA) and amino acid conjugation, thus controlling auxin homeostasis. To date, many GH3 gene families have been identified from different plant species. However, the GH3 gene family in kiwifruit (Actinidia chinensis) has not been reported. In this study, 12 AcGH3 genes were identified, phylogenetic analysis of AtGH3 (Arabidopsis), SlGH3 (Solanum lycopersicum), and AcGH3 provided insights into various orthologous relationships among these proteins, which were categorized into three groups. Expression analysis of AcGH3 genes at different postharvest stages suggested limited or no role for most of the AcGH3 genes at the initiation of fruit ripening. AcGH3.1 was the only gene exhibiting ripening-associated expression. Further study showed that the expression of AcGH3.1 gene was induced by NAA (1-naphthylacetic acid, auxin analogue) and inhibited by 1-MCP (1-methylcyclopropene, ethylene receptor inhibitor), respectively. AcGH3.1 gene silencing inhibited gene expression and delayed fruit softening in kiwifruit. The results indicate that AcGH3.1 may play an important role in the softening process of fruits. Analysis of the AcGH3.1 promoter revealed the presence of many cis-elements related to hormones, light, and drought. The determination of GUS (β-Galactosidase) enzyme activity revealed that promoter activity increased strikingly upon abscisic acid (ABA), ethylene, or NAA treatment, and significantly decreased with salicylic acid (SA) treatment. The present study could help in the identification of GH3 genes and revelation of AcGH3.1 gene function during postharvest stages, which pave the way for further functional verification of the AcGH3.1 gene.

The auxin-insensitive bodenlos mutation affects primary root formation and apical-basal patterning in the Arabidopsis embryo

In Arabidopsis embryogenesis, the primary root meristem originates from descendants of both the apical and the basal daughter cell of the zygote. We have isolated a mutant of a new gene named BODENLOS (BDL) in which the primary root meristem is not formed whereas post-embryonic roots develop and bdl seedlings give rise to fertile adult plants. Some bdl seedlings lacked not only the root but also the hypocotyl, thus resembling monopteros (mp) seedlings. In addition, bdl seedlings were insensitive to the auxin analogue 2,4-D, as determined by comparison with auxin resistant1 (axr1) seedlings. bdl embryos deviated from normal development as early as the two-cell stage at which the apical daughter cell of the zygote had divided horizontally instead of vertically. Subsequently, the uppermost derivative of the basal daughter cell, which is normally destined to become the hypophysis, divided abnormally and failed to generate the quiescent centre of the root meristem and the central root cap. We also analysed double mutants. bdl mp embryos closely resembled the two single mutants, bdl and mp, at early stages, while bdl mp seedlings essentially consisted of hypocotyl but did form primary leaves. bdl axr1 embryos approached the mp phenotype at later stages, and bdl axr1 seedlings resembled mp seedlings. Our results suggest that BDL is involved in auxin-mediated processes of apical-basal patterning in the Arabidopsis embryo.

The chemistry of 2,1-benzisothiazoles. IX. The reaction of 3-chloro-substituted 2,1-benzisothiazoles with nucleophiles and the preparation of(2,1-Benzisothiazol-3-yl)acetic acid

3-Chloro-2,1-benzisothiazole (2; R = H, R? = Cl) and 3,5-dichloro-2,1- benzisothiazole (2;R,R? = Cl) react readily with nucleophiles, and yield 3-substituted products. Acid hydrolysis of one such product, ethyl (2,1-benzisothiazol-3-yl)cyanoacetate (2; R = H, R? = CH(CN)CO2Et),affords (2,1-benzisothiazol-3-yl)acetic acid (2; R = H, R? = CH2CO2H), an auxin analogue.