Physiological transgene regulation and functional complementation of a neurological disease gene deficiency in neurons.
The microtubule-associated protein tau (MAPT) and alpha-synuclein (SNCA) genes play central roles in neurodegenerative disorders. Mutations in each gene cause familial disease, whereas common genetic variation at both loci contributes to susceptibility to sporadic neurodegenerative disease. Here, we demonstrate exquisite gene regulation of the human MAPT and SNCA transgene loci and functional complementation in neuronal cell cultures and organotypic brain slices using the herpes simplex virus type 1 (HSV-1) amplicon-based infectious bacterial artificial chromosome (iBAC) vector to express complete loci >100 kb. Cell cultures transduced by iBAC vectors carrying a 143 kb MAPT or 135 kb SNCA locus expressed the human loci similar to the endogenous gene. We focused on analysis of the iBAC-MAPT vector carrying the complete MAPT locus. On transduction into neuronal cultures, multiple MAPT transcripts were expressed from iBAC-MAPT under strict developmental and cell type-specific control. In primary neurons from Mapt(-/-) mice, the iBAC-MAPT vector expressed the human tau protein, as detected by enzyme-linked immunosorbent assay and immunocytochemistry, and restored sensitivity of Mapt(-/-) neurons to Abeta peptide treatment in dissociated neuronal cultures and in organotypic slice cultures. The faithful retention of gene expression and phenotype complementation by the system provides a novel method to analyze neurological disease genes.
Peruzzi
Pier Paolo
PP
Department of Neurological Surgery, Dardinger Laboratory for Neuro-oncology and Neurosciences, The Ohio State University Medical School, Columbus, Ohio, USA.
Lawler
Sean E
SE
Senior
Steve L
SL
Dmitrieva
Nina
N
Edser
Pauline A H
PA
Gianni
Davide
D
Chiocca
E Antonio
EA
Wade-Martins
Richard
R
D000818:Animals; D002478:Cells, Cultured; D022202:Chromosomes, Artificial, Bacterial; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D007150:Immunohistochemistry; D051379:Mice; D009422:Nervous System Diseases / Q000235:genetics / Q000378:metabolism / Q000628:therapy*; D009474:Neurons / Q000378:metabolism / Q000473:pathology*; D051381:Rats; D019076:Transgenes / Q000235:genetics* / Q000502:physiology*; D051844:alpha-Synuclein / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D016875:tau Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology
C054369:MAPT protein, human; D051844:alpha-Synuclein; D016875:tau Proteins