The researchers hit upon the sugar as a candidate after they had addressed the difficulty of getting poly-Q proteins such as huntingtin (htt) into solution. This step was necessary because the authors wanted to search for aggregation inhibitors with high-throughput in-vitro screening. As a surrogate for disease-causing poly-Q proteins, the researchers employed a mutant sperm whale myoglobin carrying glutamine repeats. This protein mimicked the native poly-Q proteins and could be dissolved, allowing the authors to screen a number of small, nontoxic molecules that can administered orally. Various disaccharides proved to inhibit aggregation in vitro and also correlated with decreased cell death in cultured neuroblastoma cells transfected with an N-terminal huntingtin fragment.

Trehalose, the most effective aggregation inhibitor in these experiments, was then tested in the R6/2 mouse model of HD. Mice that had been ingesting the disaccharide in their water from about three weeks of age showed differences in brain pathology from their untreated counterparts by 12 weeks of age. The dilation of the ventricles seen in this HD model was significantly reduced, as was the number of ubiquitin-positive inclusions in motor cortex, striatum, and liver. The mice showed improved motor performance on the rotorod and in footprinting tests, and lived longer.

How could this sugar work? Cell-stress pathways, for one, do not appear to mediate its effects. "Instead, the inhibitory effects of trehalose on the aggregation of two different polyglutamine-bearing proteins—Mb-Gln35 and truncated huntingtin—indicate that trehalose may bind directly to expanded polyglutamines," write the authors.

While this simple sugar proves itself as a treatment for Huntington's patients—or fails as such—research continues apace to define the mechanisms by which huntingtin induces neurodegeneration. In the January 7 Journal of Neuroscience, a team led by Zheng-Hong Qin and Marian DiFiglia of Massachusetts General Hospital in Charlestown report on their in-vitro investigations of how mutant huntingtin protein may sequester other proteins. DiFiglia's research group has previously reported on curious multivesicular bodies that form from the fusion of autophagosomes in cultured cells expressing wild-type or mutant htt (Kegel et al., 2000). There is also evidence, though not conclusive, that such structures, dubbed "htt bodies" by the authors, are found in neurons in Huntington's disease (Sapp et al., 1997).

In the current study, the researchers examined these htt bodies more closely and found that poly-Q expansion increased their formation. Within the fused autophagosomes, some of the mutant proteins—those that had congregated at the periphery of the bodies in oligomeric globules—appear to have entrapped a number of cytoplasmic proteins typically found elsewhere in the cell. Among these were heat shock protein 70, proteasome, dynamin, htt-interacting protein 1 (HIP1), SH3-containing Grb2-like protein (SH3GL3), and 14.7K-interacting protein. By contrast, the mutant htt found in the core of the vesicles was fibrillar and protease-resistant, and associated with cathepsin D, ubiquitin, and heat shock protein 40.

Adjacent to the poly-Q region in htt is a polyproline region that might interact with proteins with Src homology 3 (SH3) and WW domains; this might add structural stability to htt, the authors speculate. Removing this region led to a reduction in htt bodies and blocked the sequestering of proteins in the shell.

All in all, this evidence supports the idea that vesicle trafficking is perturbed in HD. Testing this idea directly, the authors found that the endocytic uptake of transferrin was significantly reduced in cells with htt bodies. Finally, the authors looked for evidence of endocytic perturbations in human tissue. In cortical neurons of HD patients with early pathology, they found that the distribution of dynamin already was altered.

"Our findings suggest that soluble oligomers of mutant htt that assemble at the periphery of htt bodies may cause cell dysfunction in HD by sequestering proteins involved in vesicle trafficking through a polyproline-dependent mechanism," conclude the authors.—Hakon Heimer.

References:
Tanaka M, Machida Y, Niu S, Ikeda T, Jana NR, Doi H, Kurosawa M, Nekooki M, Nukina N. Trehalose alleviates polyglutamine-mediated pathology in a mouse model of Huntington disease. Nat Med. 2004 Jan 18 [Epub ahead of print]. Abstract

Qin ZH, Wang Y, Sapp E, Cuiffo B, Wanker E, Hayden MR, Kegel KB, Aronin N, DiFiglia M. Huntingtin bodies sequester vesicle-associated proteins by a polyproline-dependent interaction. J Neurosci. 2004 Jan 7; 24(1): 269-81. Abstract