Multiple Roles of the Low-Affinity Calcium Uptake System in Drechslerella dactyloides, a Nematode-Trapping Fungus That Forms Constricting Rings.

(1) Background: the low-affinity calcium uptake system (LACS) has been shown to play a crucial role in the conidiation and formation of adhesive nets and knobs by nematode-trapping fungi (NTF), but its involvement in the formation of constricting rings (CRs), mechanical traps to capture free-living nematodes, remains unexplored. (2) Methods: we investigated the function of two LACS genes (DdaFIG_1 and DdaFIG_2) in Drechslerella dactyloides, an NTF that forms CRs. We generated single (DdaFIG_1Ri and DdaFIG_2Ri) and double (DdaFIG_1,2Ri) knockdown mutants via the use of RNA interference (RNAi). (3) Results: suppression of these genes significantly affected conidiation, trap formation, vegetative growth, and response to diverse abiotic stresses. The number of CRs formed by DdaFIG_1Ri, DdaFIG_2Ri, and DdaFIG_1,2Ri decreased to 58.5%, 59.1%, and 38.9% of the wild-type (WT) level, respectively. The ring cell inflation rate also decreased to 73.6%, 60.6%, and 48.8% of the WT level, respectively. (4) Conclusions: the LACS plays multiple critical roles in diverse NTF.

Characterization of the complete mitochondrial genome of the nematode-trapping fungus Drechslerella dactyloides.

The complete mitochondrial genome of Drechslerella dactyloides was characterized in this study. This mitogenome is a closed circular molecule of 246860 bp in length with a GC content of 26.16%, including 87 predicted protein-coding genes, 29 transfer RNA genes, and two rRNA gens. Phylogenetic analyses based on concatenated amino acid sequences at 14 conserved mitochondrial protein-coding genes showed that D. dactyloides was closely related to Dactylellina haptotyla.

SNARE Protein DdVam7 of the Nematode-Trapping Fungus Drechslerella dactyloides Regulates Vegetative Growth, Conidiation, and the Predatory Process via Vacuole Assembly.

Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins play conserved roles in membrane fusion events in eukaryotes and have been documented to be involved in fungal growth and pathogenesis. However, little is known about the roles of SNAREs in trap morphogenesis in nematode-trapping fungi (NTF). Drechslerella dactyloides, one of the constricting ring-forming NTF, captures free-living nematodes via rapid ring cell inflation. Here, we characterized DdVam7 of D. dactyloides, a homolog of the yeast SNARE protein Vam7p. Deletion of DdVam7 significantly suppressed vegetative growth and conidiation. The mutation significantly impaired trap formation and ring cell inflation, resulting in a markedly decreased nematode-trapping ability. A large vacuole could develop in ring cells within ~2.5 s after instant inflation in D. dactyloides. In the ΔDdVam7 mutant, the vacuoles were small and fragmented in hyphae and uninflated ring cells, and the large vacuole failed to form in inflated ring cells. The localization of DdVam7 in vacuoles suggests its involvement in vacuole fusion. In summary, our results suggest that DdVam7 regulates vegetative growth, conidiation, and the predatory process by mediating vacuole assembly in D. dactyloides, and this provides a basis for studying mechanisms of SNAREs in NTF and ring cell rapid inflation. IMPORTANCE D. dactyloides is a nematode-trapping fungus that can capture nematodes through a constricting ring, the most sophisticated trapping device. It is amazing that constricting ring cells can inflate to triple their size within seconds to capture a nematode. A large centrally located vacuole is a unique signature associated with inflated ring cells. However, the mechanism underpinning trap morphogenesis, especially vacuole dynamics during ring cell inflation, remains unclear. Here, we documented the dynamics of vacuole assembly during ring cell inflation via time-lapse imaging for the first time. We characterized a SNARE protein in D. dactyloides (DdVam7) that was involved in vacuole assembly in hyphae and ring cells and played important roles in vegetative growth, conidiation, trap morphogenesis, and ring cell inflation. Overall, this study expands our understanding of biological functions of the SNARE proteins and vacuole assembly in NTF trap morphogenesis and provides a foundation for further study of ring cell rapid inflation mechanisms.

DdaCrz1, a C2H2-Type Transcription Factor, Regulates Growth, Conidiation, and Stress Resistance in the Nematode-Trapping Fungus Drechslerella dactyloides.

The Ca2+/calmodulin-dependent signaling pathway regulates diverse cellular processes. Calcineurin is a calcium-dependent phosphatase acting in fungi mainly through Crz1, a zinc finger transcription factor. Although the likely involvement of Ca2+ in fungal carnivorism has been documented, how Crz1 functions in nematode-trapping fungi remains unknown. Here, we identified the Crz1 gene (named as DdaCrz1) in Drechslerella dactyloides, a species that forms constricting rings to trap nematodes. The deletion of DdaCrz1 significantly reduced hyphal growth and conidiation, trap formation, and ring cell inflation. Moreover, the mutation increased sensitivity to Mn2+ but decreased sensitivity to Ca2+, Mg2+, Zn2+, and Li+. Similarly, the mutant showed increased tolerance to osmotic stress but was more sensitive to Congo red, a cell wall-damaging agent. Our results confirmed the critical roles of the Ca2+/calmodulin-dependent signaling pathway in regulating growth, conidiation, and the stress response, and suggested its involvement in trapping nematodes.