Evaluation of multi-color genetically encoded Ca2+ indicators in filamentous fungi.

Evaluation of multi-color genetically encoded Ca2+ indicators in filamentous fungi. Fungal Genet Biol. 2021 Feb 16;:103540 Authors: Kim HS, Kim JE, Hwangbo A, Akerboom J, Looger LL, Duncan R, Son H, Czymmek KJ, Kang S Abstract Genetically encoded Ca2+ indicators (GECIs) enable long-term monitoring of cellular and subcellular dynamics of this second messenger in response to environmental and developmental cues without relying on exogenous dyes. Continued development and optimization in GECIs, combined with advances in gene manipulation, offer new opportunities for investigating the mechanism of Ca2+ signaling in fungi, ranging from documenting Ca2+ signatures under diverse conditions and genetic backgrounds to evaluating how changes in Ca2+ signature impact calcium-binding proteins and subsequent cellular changes. Here, we attempted to express multi-color (green, yellow, blue, cyan, and red) circularly permuted fluorescent protein (FP)-based Ca2+ indicators driven by multiple fungal promoters in Fusarium oxysporum, F. graminearum, and Neurospora crassa. Several variants were successfully expressed, with GCaMP5G driven by the Magnaporthe oryzae ribosomal protein 27 (PRP27) and F. verticillioides elongation factor-1α (PEF1α) gene promoters being optimal for F. graminearum and F. oxysporum, respectively. Transformants expressing GCaMP5G were compared with those expressing YC3.60, a ratiometric Cameleon Ca2+ indicator. Wild-type and thr...
Source: Fungal Genetics and Biology - Category: Biology Authors: Tags: Fungal Genet Biol Source Type: research