Heterologous expression of a green fluorescent protein–pertussis toxin S1 subunit fusion construct disrupts calcium channel modulation in rat superior cervical ganglion neurons

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• 作者：Ikeda ; Stephen R. ; Jeong ; Seong-Woo ; Kammermeier ; Paul J. ; Ruiz-Velasco ; Victor ; King ; Marina M.
• 关键词：Sympathetic neurons ; Pertussis toxin ; Green fluorescent protein ; G-protein ; Calcium channels ; Potassium channels
• 刊名：Neuroscience Letters
• 出版年：1999
• 出版时间：August 27, 1999
• 年：1999
• 卷：271
• 期：3
• 页码：179-182
• 全文大小：124 K

文摘

The fusion construct pEGFP-PTXS1 was assembled by ligating cDNA encoding the S1 subunit of Bordetella pertussis toxin (PTX) into the plasmid pEGFP-C1 (which codes for enhanced green fluorescent protein). Microinjection of pEGFP-PTXS1 (1–100 ng/μl) into the nucleus of dissociated rat sympathetic ganglion neurons resulted in functional expression as determined from the diffuse green fluorescence and disruption of norepinephrine-mediated N-type Ca²⁺ channel modulation. The heterologously expressed toxin retained specificity for Gα_i/o-dependent pathways as VIP-mediated modulation of N-type Ca²⁺ channels and muscarine-mediated inhibition of M-type K⁺ channels persisted in pEGFP-PTXS1 expressing neurons. These data demonstrate that the S1 subunit of PTX is readily expressed in mammalian neurons and remains functional following fusion to the C-terminus of another protein.