Targeted ablation of beta cells in the embryonic zebrafish pancreas using E. coli nitroreductase.
In order to generate a zebrafish model of beta cell regeneration, we have expressed an Escherichia coli gene called nfsB in the beta cells of embryonic zebrafish. This bacterial gene encodes a nitroreductase (NTR) enzyme, which can convert prodrugs such as metronidazole (Met) to cytotoxins. By fusing nfsB to mCherry, we can simultaneously render beta cells susceptible to prodrug and visualize Met dependent cell ablation. We show that the neighboring alpha and delta cells are unaffected by prodrug treatment and that ablation is beta cell specific. Following drug removal and 36h of recovery, beta cells regenerate. Using ptf1a morphants, it is clear that this beta cell recovery occurs independently of the presence of the exocrine pancreas. Also, by using photoconvertible Kaede to cell lineage trace and BrdU incorporation to label proliferation, we investigate mechanisms for beta regeneration. Therefore, we have developed a unique resource for the study of beta cell regeneration in a living vertebrate organism, which will provide the opportunity to conduct large-scale screens for pharmacological and genetic modifiers of beta cell regeneration.
Pisharath
Harshan
H
Department of Comparative Medicine, Johns Hopkins School of Medicine, 720 Rutland Avenue, Ross 771, Baltimore, MD 21205, USA.
Rhee
Jerry M
JM
Swanson
Michelle A
MA
Leach
Steven D
SD
Parsons
Michael J
MJ
D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D003920:Diabetes Mellitus; D004195:Disease Models, Animal; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000502:physiology*; D050417:Insulin-Secreting Cells / Q000378:metabolism* / Q000473:pathology; D009601:Nitroreductases / Q000502:physiology*; D010179:Pancreas / Q000166:cytology / Q000196:embryology / Q000201:enzymology* / Q000473:pathology; D012038:Regeneration / Q000235:genetics; D015027:Zebrafish / Q000196:embryology*
D029968:Escherichia coli Proteins; C116053:NfsA protein, E coli; D009601:Nitroreductases