Optimization of adeno-associated viral vector-mediated transduction of the corticospinal tract: comparison of four promoters

Bart Nieuwenhuis,Barbara Hobo,Joost Verhaagen,Sam Hilton,Barbara Haenzi,Alejandro Carnicer-Lombarte,James W Fawcett

doi:10.1038/s41434-020-0169-1

Abstract

Adeno-associated viral vectors are widely used as vehicles for gene transfer to the nervous system. The promoter and viral vector serotype are two key factors that determine the expression dynamics of the transgene. A previous comparative study has demonstrated that AAV1 displays efficient transduction of layer V corticospinal neurons, but the optimal promoter for transgene expression in corticospinal neurons has not been determined yet. In this paper, we report a side-by-side comparison between four commonly used promoters: the short CMV early enhancer/chicken β actin (sCAG), human cytomegalovirus (hCMV), mouse phosphoglycerate kinase (mPGK) and human synapsin (hSYN) promoter. Reporter constructs with each of these promoters were packaged in AAV1, and were injected in the sensorimotor cortex of rats and mice in order to transduce the corticospinal tract. Transgene expression levels and the cellular transduction profile were examined after 6 weeks. The AAV1 vectors harbouring the hCMV and sCAG promoters resulted in transgene expression in neurons, astrocytes and oligodendrocytes. The mPGK and hSYN promoters directed the strongest transgene expression. The mPGK promoter did drive expression in cortical neurons and oligodendrocytes, while transduction with AAV harbouring the hSYN promoter resulted in neuron-specific expression, including perineuronal net expressing interneurons and layer V corticospinal neurons. This promoter comparison study contributes to improve transgene delivery into the brain and spinal cord. The optimized transduction of the corticospinal tract will be beneficial for spinal cord injury research.

Highlights

Supplementary information The online version of this article contains supplementary material, which is available to authorized users.The corticospinal tract (CST) is an important descending motor pathway controlling the movement of the limbs and trunk
Four days after the transfection, the eGFP intensity was similar between the sCAG, human cytomegalovirus (hCMV), mouse phosphoglycerate kinase (mPGK) and human synapsin (hSYN) promoters (Fig. 3b, c)
Consistent with that study, we found in both mice and rats that AAV1 transduces almost twice as many cortical neurons and layer V corticospinal neurons as AAV5 (Supplementary Figs. 2, 3), highlighting that AAV1 is superior to AAV5 for transduction of the CST and cortex

Summary

Introduction

The corticospinal tract (CST) is an important descending motor pathway controlling the movement of the limbs and trunk. Damage to the CST results in paralysis. Repair of the CST is limited because this motor pathway has a low intrinsic neuronal capacity for axon regeneration. The axon regeneration capacity of corticospinal neurons can be enhanced by delivering regeneration-associated genes using gene therapy (reviewed in [1,2,3,4]). Manipulation of the CST of rats and mice is widely used to examine the neuronintrinsic mechanisms that underlie the failure of axon regeneration

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