An Improved Mesocotyl Elongation Assay for the Rapid Identification and Characterization of Strigolactone-Related Rice Mutants

Strigolactones (SLs) constitute an important class of plant hormones involved in diverse developmental activities in plant growth and host-parasite interaction. Although substantial progress has been made to understand this pathway, the mechanism of action is still elusive especially with its interaction with other phytohormones and downstream targets. Here we have utilized the negative role of strigolactones in rice (Oryza sativa L.) mesocotyl elongation as a morphological marker for the identification and characterization of new developmental mutants. We observed that deep sown seeds develop longer mesocotyl compared with the surface-grown seeds in the dark condition. Based on this observation, we have developed a method to access mesocotyl elongation consisting of the glass vessel and vermiculite as a growth media. Mesocotyl elongation in the modified deep sown system results in a many-fold increase compared to the surface-grown seeds in the dark condition. External application of SLs analog rac-GR24 rescued the elongated mesocotyl phenotype in the mutant defective in SLs synthesis but not the signaling mutant, demonstrating its applicability in the physiological experiments. The modified mesocotyl elongation assay can be used as a rapid method for characterization and identification of suppressors/enhancers and new developmental mutants in the SLs or its associated pathway saving a huge amount of time and space.

Submerged Conidiation and Product Formation by Aspergillus niger at Low Specific Growth Rates Are Affected in Aerial Developmental Mutants

ABSTRACTExposure to an aerial environment or severe nutrient limitation induces asexual differentiation in filamentous fungi. Submerged cultivation ofAspergillus nigerin carbon- and energy-limited retentostat cultures both induces and fuels conidiation. Physiological and transcriptomic analyses have revealed that this differentiation strongly affects product formation. Since conidiation is inherent in the aerial environment, we hypothesized that product formation near zero growth can be influenced by affecting differentiation or development of aerial hyphae in general. To investigate this idea, three developmental mutants (ΔfwnA,scl-1, andscl-2mutants) that have no apparent vegetative growth defects were cultured in maltose-limited retentostat cultures. The secondary-metabolite profile of the wild-type strain defined flavasperone, aurasperone B, tensidol B, and two so far uncharacterized compounds as associated with conidium formation, while fumonisins B2, B4, and B6were characteristic of early response to nutrient limitation by the vegetative mycelium. The developmental mutants responded differently to the severe substrate limitation, which resulted in distinct profiles of growth and product formation.fwnAencodes the polyketide synthase responsible for melanin biosynthesis during aerial differentiation, and we show that conidial melanin synthesis in submerged retentostat cultures and aurasperone B production arefwnAdependent. Thescl-1andscl-2strains are two UV mutants generated in the ΔfwnAbackground that displayed reduced asexual conidiation and formed sclerotium-like structures on agar plates. The reduced conidiation phenotypes of thescl-1andscl-2strains are reflected in the retentostat cultivation and are accompanied by elimination or severely reduced accumulation of secondary metabolites and distinctly enhanced accumulation of extracellular protein. This investigation shows that submerged conidiation and product formation of a mitosporic fungus cultured at low specific growth rates can be fundamentally affected by interfering with the genetic program for differentiation of aerial hyphae, opening new perspectives for tailoring industrial performance.