Calcium measurements in living filamentous fungi expressing codon-optimized aequorin

G. Nelson; O. Kozlova-Zwinderman; A. J. Collis; M. R. Knight; J. R S Fincham; C. P. Stanger; A. Renwick; J. G M Hessing; P. J. Punt; C. A M J J Van Den Hondel; N. D. Read

doi:10.1111/j.1365-2958.2004.04066.x

Calcium measurements in living filamentous fungi expressing codon-optimized aequorin

G. Nelson
, O. Kozlova-Zwinderman
, A. J. Collis
, M. R. Knight
, J. R S Fincham
, C. P. Stanger
, A. Renwick
, J. G M Hessing
, P. J. Punt
, C. A M J J Van Den Hondel
, N. D. Read

Research output: Contribution to journal › Article › peer-review

Abstract

Calcium signalling is little understood in filamentous fungi largely because easy and routine methods for calcium measurement in living hyphae have previously been unavailable. We have developed the recombinant aequorin method for this purpose. High levels of aequorin expression were obtained in Neurospora crassa, Aspergillus niger and Aspergillus awamori by codon optimization of the aequorin gene. Three external stimuli (mechanical perturbation, hypoosmotic shock and high external calcium) were found transiently to increase [Ca 2+]c. Each of the calcium signatures associated with these physico-chemical treatments was unique, suggesting the involvement of three distinct calcium-mediated signal transduction pathways. The fungal calcium channel blocker KP4 inhibited the [Ca2+]c responses to hypo-osmotic shock and high external calcium, but not to mechanical perturbation. The divalent cation chelator BAPTA inhibited [Ca 2+]c responses to mechanical perturbation and hypo-osmotic shock. The calcium agonists A23187 and cyclopiazonic acid increased [Ca 2+]c levels.

Original language	English
Pages (from-to)	1437-1450
Number of pages	13
Journal	Molecular Microbiology
Volume	52
Issue number	5
DOIs	https://doi.org/10.1111/j.1365-2958.2004.04066.x
Publication status	Published - Jun 2004

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Nelson, G., Kozlova-Zwinderman, O., Collis, A. J., Knight, M. R., Fincham, J. R. S., Stanger, C. P., Renwick, A., Hessing, J. G. M., Punt, P. J., Van Den Hondel, C. A. M. J. J., & Read, N. D. (2004). Calcium measurements in living filamentous fungi expressing codon-optimized aequorin. Molecular Microbiology, 52(5), 1437-1450. https://doi.org/10.1111/j.1365-2958.2004.04066.x

@article{2ece782103cd469e8fdb0e1595667b38,

title = "Calcium measurements in living filamentous fungi expressing codon-optimized aequorin",

abstract = "Calcium signalling is little understood in filamentous fungi largely because easy and routine methods for calcium measurement in living hyphae have previously been unavailable. We have developed the recombinant aequorin method for this purpose. High levels of aequorin expression were obtained in Neurospora crassa, Aspergillus niger and Aspergillus awamori by codon optimization of the aequorin gene. Three external stimuli (mechanical perturbation, hypoosmotic shock and high external calcium) were found transiently to increase [Ca 2+]c. Each of the calcium signatures associated with these physico-chemical treatments was unique, suggesting the involvement of three distinct calcium-mediated signal transduction pathways. The fungal calcium channel blocker KP4 inhibited the [Ca2+]c responses to hypo-osmotic shock and high external calcium, but not to mechanical perturbation. The divalent cation chelator BAPTA inhibited [Ca 2+]c responses to mechanical perturbation and hypo-osmotic shock. The calcium agonists A23187 and cyclopiazonic acid increased [Ca 2+]c levels.",

author = "G. Nelson and O. Kozlova-Zwinderman and Collis, \{A. J.\} and Knight, \{M. R.\} and Fincham, \{J. R S\} and Stanger, \{C. P.\} and A. Renwick and Hessing, \{J. G M\} and Punt, \{P. J.\} and \{Van Den Hondel\}, \{C. A M J J\} and Read, \{N. D.\}",

year = "2004",

month = jun,

doi = "10.1111/j.1365-2958.2004.04066.x",

language = "English",

volume = "52",

pages = "1437--1450",

journal = "Molecular Microbiology",

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T1 - Calcium measurements in living filamentous fungi expressing codon-optimized aequorin

AU - Nelson, G.

AU - Kozlova-Zwinderman, O.

AU - Collis, A. J.

AU - Knight, M. R.

AU - Fincham, J. R S

AU - Stanger, C. P.

AU - Renwick, A.

AU - Hessing, J. G M

AU - Punt, P. J.

AU - Van Den Hondel, C. A M J J

AU - Read, N. D.

PY - 2004/6

Y1 - 2004/6

N2 - Calcium signalling is little understood in filamentous fungi largely because easy and routine methods for calcium measurement in living hyphae have previously been unavailable. We have developed the recombinant aequorin method for this purpose. High levels of aequorin expression were obtained in Neurospora crassa, Aspergillus niger and Aspergillus awamori by codon optimization of the aequorin gene. Three external stimuli (mechanical perturbation, hypoosmotic shock and high external calcium) were found transiently to increase [Ca 2+]c. Each of the calcium signatures associated with these physico-chemical treatments was unique, suggesting the involvement of three distinct calcium-mediated signal transduction pathways. The fungal calcium channel blocker KP4 inhibited the [Ca2+]c responses to hypo-osmotic shock and high external calcium, but not to mechanical perturbation. The divalent cation chelator BAPTA inhibited [Ca 2+]c responses to mechanical perturbation and hypo-osmotic shock. The calcium agonists A23187 and cyclopiazonic acid increased [Ca 2+]c levels.

AB - Calcium signalling is little understood in filamentous fungi largely because easy and routine methods for calcium measurement in living hyphae have previously been unavailable. We have developed the recombinant aequorin method for this purpose. High levels of aequorin expression were obtained in Neurospora crassa, Aspergillus niger and Aspergillus awamori by codon optimization of the aequorin gene. Three external stimuli (mechanical perturbation, hypoosmotic shock and high external calcium) were found transiently to increase [Ca 2+]c. Each of the calcium signatures associated with these physico-chemical treatments was unique, suggesting the involvement of three distinct calcium-mediated signal transduction pathways. The fungal calcium channel blocker KP4 inhibited the [Ca2+]c responses to hypo-osmotic shock and high external calcium, but not to mechanical perturbation. The divalent cation chelator BAPTA inhibited [Ca 2+]c responses to mechanical perturbation and hypo-osmotic shock. The calcium agonists A23187 and cyclopiazonic acid increased [Ca 2+]c levels.

U2 - 10.1111/j.1365-2958.2004.04066.x

DO - 10.1111/j.1365-2958.2004.04066.x

M3 - Article

C2 - 15165245

VL - 52

SP - 1437

EP - 1450

JO - Molecular Microbiology

JF - Molecular Microbiology

IS - 5

ER -

Calcium measurements in living filamentous fungi expressing codon-optimized aequorin

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