1 December 2006. The NAD synthesizing protein nicotinamide mononucleotide adenylyltransferase (Nmnat) got the attention of neuroscience researchers as one-half of a chimeric, neuroprotective protein responsible for slowing Wallerian degeneration in the Wlds mutant mouse (see ARF related news story). It was considered the lesser half, until experiments showed that overexpression of just the Nmnat enzyme was sufficient to protect axons from neurodegeneration (see ARF related news story). Now some new data secure Nmnat’s standing as a bona fide neuroprotective protein, but not precisely as expected.
In a paper published in the December issue of PLoS Biology, Hugo Bellen and colleagues at Baylor College of Medicine in Houston, Texas, demonstrate that Nmnat is crucial for saving photoreceptor neurons from activity-induced degeneration in Drosophila. The surprising finding is that, contrary to previous suggestions, the protective power of fly Nmnat does not rely on NAD synthesis activity.
The researchers, led by first author Grace Zhai and colleagues, isolated the Drosophila Nmnat gene in a forward genetic screen for synapse malfunction. They used mosaic flies that carry homozygous mutations restricted to the visual system, a strategy that allows for the isolation of mutants that would be lethal if absent completely from the animals. They first screened for loss of phototaxis, and then looked for mutants with normal gross eye morphology but abnormal photoreceptor axons, terminals, and synaptic function.
Two blind mutants with poor synapse structure and function turned out to have substitutions in the coding region of a gene that the researchers identified as the Drosophila version of human Nmnat. The mutations, which created null alleles, caused an age-dependent neurodegeneration of photoreceptor cells. Eye development appeared normal, and the degeneration was activity-dependent: rearing flies in the dark, or interrupting the phototransduction cascade with other mutations, retarded neuron loss.
To test if the phenotype was due to loss of NAD synthase activity, the researchers attempted to rescue the neurodegeneration using different Nmnat constructs. They found that two mutants, each one showing a 99 percent reduction in enzymatic activity, were just as effective at supporting neuronal survival as a wild-type human NMNAT3. However, the mutants could not reverse the lethality of a ubiquitous Nmnat homozygous deletion, indicating that the protein has at least two independent roles—one required for organismal viability and another for neuroprotection.
In the Wlds mouse, overexpression of the Nmnat fusion protein delays degeneration, and the researchers saw a similar effect after overexpression of Nmnat in flies. Introduction of either wild-type or mutant, catalytically inactive protein prevented degeneration due to two different retinal degeneration mutants, and also in response to excessive neuronal activity induced by constant intense light.
The demonstration that catalytically inactive Nmnat is just as effective at neuroprotection is at odds with earlier reports on overexpression of the proteins in vitro (see ARF related news story), which showed that enzymatic activity and NAD were required to protect axon degeneration. While the reasons for this discrepancy are not clear, the current results are consistent with findings in the original Wlds mice in that even as the levels of Nmnat were increased thanks to the chimera, NAD concentrations were not.
The study puts the question of how Nmnat protects neurons back to square one. But the authors suggest the protein might be “exploited to protect neurons against activity-induced neurodegeneration,” a prospect that might be of some use in Alzheimer and other neurodegenerative diseases.—Pat McCaffrey.
Zhai RG, Cao Y, Hiesinger PR, Zhou Y, Mehta SQ, Schulze KL, Verstreken P, Bellen HJ. Drosophila NMNAT Maintains Neural Integrity Independent of Its NAD Synthesis Activity. PLoS Biology. 2006 November 28. Abstract