The T alpha 1 alpha-tubulin promoter specifies gene expression as a function of neuronal growth and regeneration in transgenic mice

Abstract

We have previously demonstrated that one member of the alpha-tubulin multigene family, termed T alpha 1 in rats, is regulated as a function of neuronal growth and regeneration. To elucidate the molecular mechanisms responsible for coupling gene expression to morphological differentiation, we have isolated the T alpha 1 gene, have fused 1.1 kb of the 5' flanking region to a nuclear lacZ reporter gene, and have generated transgenic mice. Analysis of these transgenic mice demonstrated that marker gene expression was specific to the CNS and PNS, with expression in vivo at embryonic day 13.5 being similar to expression of the endogenous gene. Moreover, the induction of transgene expression was correlated temporally with neuronal commitment in developing neural crest-derived peripheral neurons and in the developing retina. Immunocytochemical analysis of mixed primary embryonic brain cultures confirmed that transgene expression was specific to neurons, with the majority of neurons, but not astrocytes or oligodendrocytes, expressing beta-galactosidase. Transgene expression in vivo was maintained in developing neurons until early in postnatal life, subsequent to which its expression decreased coincident with neuronal maturation. The transgene was then reinduced in regenerating facial motoneurons following unilateral axotomy of the facial nerve. Thus, 1.1 kb of 5' flanking sequence from the T alpha 1 gene contains the sequence elements responsible for specifying gene expression to embryonic neurons and for subsequently regulating gene expression in both developing and mature neurons as a function of morphological growth.

Neuronal expression of the transcription factor Gli1 using the Tα1 α-tubulin promoter is neuroprotective in an experimental model of Parkinson's disease

“…37, 38 We constructed first-generation RAd vectors expressing Gli1 or the control marker gene lacZ with a nuclear localization signal. A diagram illustrating the construction of the RAds and the molecular analysis of the recombinant genomes is shown in Figure 1.…”

“…We utilized the Ta1 promoter element that has been shown to mediate neuronal-specific expression in transgenic animals and in neurons in vitro. 37,38,45,46 To determine whether the Ta1 promoter would retain its neuronal-specific pattern of expression within the context of an adenoviral vector, we constructed a recombinant adenoviral vector where Ta1 was cloned upstream of the gene encoding b-galactosidase containing a nuclear localization signal. Double labeling of neurons following the injection of the Ta1-lacZ vector into the striatum indicated that more than 95% of cells transduced and expressing the reporter gene also expressed the neuronal marker Neu-N indicating the neuronal phenotype of transduced cells.…”

Mistargeting hippocampal axons by expression of a truncated Eph receptor

“…The fusion protein contains the extracellular ligand-binding and transmembrane domains of mouse EphA5, as well as the GFP coding sequence. The entire fusion gene was then cloned into the P253 vector kindly provided by Freda Miller (Montreal Neurological Institute, Montreal) under the transcriptional control of a neuron-specific ␣-tubulin promoter (27).…”

Efficient gene transduction of neurons by lentivirus with enhanced neuron-specific promoters

“…9 As lentiviral vectors cannot accept large genetic sequences (up to 10 kb between the long terminal repeats), we selected relatively small promoters. In the present study, we used five kinds of neuron-specific promoters; human synapsin I (SYN), 10 mouse calcium/ calmodulin-dependent protein kinase II (CaMKII), 11 rat tubulin alpha I (Ta1), 12 rat neuron-specific enolase (NSE) 13 and human platelet-derived growth factor-beta chain (PDGF) promoters. 14 Although these promoters are known to be specific for neuronal expression, the transcriptional activities are much weaker than those of viral promoters such as CMV promoter.…”