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BackgroundThe Arabidopsis microRNA156 (miR156) regulates 11 members of the SQUAMOSA PROMOTER BINDING PROTEIN LIKE (SPL) family by base pairing to complementary target mRNAs. Each SPL gene further regulates a set of other genes; thus, miR156 controls numerous genes through a complex gene regulation network. Increased axillary branching occurs in transgenic Arabidopsis overexpressing miR156b, similar to that observed in loss-of-function max3 and max4 mutants with lesions in carotenoid cleavage dioxygenases. Arabidopsis miR156b was found to enhance carotenoid levels and reproductive shoot branching when expressed in Brassica napus, suggesting a link between miR156b expression and carotenoid metabolism. However, details of the miR156 regulatory network of SPL genes related to carotenoid metabolism are not known.ResultsIn this study, an Arabidopsis T-DNA enhancer mutant, sk156, was identified due to its altered branching and trichome morphology and increased seed carotenoid levels compared to wild type (WT) ecovar Columbia. Enhanced miR156b expression due to the 35S enhancers present on the T-DNA insert was responsible for these phenotypes. Constitutive and leaf primodium-specific expression of a miR156-insensitive (mutated) SPL15 (SPL15m) largely restored WT seed carotenoid levels and plant morphology when expressed in sk156. The Arabidopsis native miR156-sensitive SPL15 (SPL15n) and SPL15m driven by a native SPL15 promoter did not restore the WT phenotype in sk156. Our findings suggest that SPL15 function is somewhat redundant with other SPL family members, which collectively affect plant phenotypes. Moreover, substantially decreased miR156b transcript levels in sk156 expressing SPL15m, together with the presence of multiple repeats of SPL-binding GTAC core sequence close to the miR156b transcription start site, suggested feedback regulation of miR156b expression by SPL15. This was supported by the demonstration of specific in vitro interaction between DNA-binding SBP domain of SPL15 and the proximal promoter sequence of miR156b.ConclusionsEnhanced miR156b expression in sk156 leads to the mutant phenotype including carotenoid levels in the seed through suppression of SPL15 and other SPL target genes. Moreover, SPL15 has a regulatory role not only for downstream components, but also for its own upstream regulator miR156b.

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Under submerged growth in a defined medium (TKI broth), the entomopathogenic fungus, Beauveria bassiana, produced conidia; it produced only blastospores in complex media. Production of such "submerged" conidia depended on the nature of the carbon source and the presence of nitrate as a nitrogen source. Maximum yield of conidia (5 × 108 mL) was obtained when glucose was the carbon source and when the glucose to nitrate ratio was 5:1. Other carbon sources gave rise to both conidia and blastospores. Reducing the phosphate concentration resulted in the production of conidia which resembled "aerial" conidia in morphology and germination rates. The surfaces of "submerged" conidia were relatively smooth, but had a tendency to acquire the rough, warty, brittle surface characteristics of aerial conidia. Blastospores produced in defined media gave rise to conidia through microcycle conidiation without going through the vegetative phase of growth. In more complex media, blastospores did not undergo microcycle conidiation.

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The surface properties of aerial conidia (AC) from 24 strains of entomopathogenic fungi were studied and compared using the salt-mediated aggregation and sedimentation (SAS) assay, electron microscopy, FITC-labelled lectins, and spore dimensions. Spores with rugose surfaces were hydrophobic, whereas hydrophilic spores had smooth surfaces. Correlation analysis found no link between spore dimensions and either hydrophobicity or surface carbohydrates. However, there was a strong positive correlation between spore hydrophobicity and surface carbohydrates. The three spore types of Beauveria bassiana were all shown to possess discrete surface hydrophobicities, which were also strongly linked to surface carbohydrate profiles. Various chemical treatments had pronounced effects on spore surface properties, with sodium dodecyl sulfate (SDS) and formic acid (FA) reducing both lectin binding and surface hydrophobicity. When FA-protein extracts were separated and analysed using SDS-PAGE, only the hydrophobic spores had low molecular weight hydrophobin-like peptides that were unglycosylated and contained disulfide bonds. The strains with hydrophilic AC had much lower levels of FA-extractable protein per spore dry weight compared to their more hydrophobic counterparts. Moreover, extracts of the more hydrophobic spores tended to have greater protein:carbohydrate ratios.Key words: fungi, spores, hydrophobicity, lectins, morphology, microbial insecticides, protein.