Expecting that degradation needs to vary depending on tissue type, age, and environmental factors, first author Marco Sardiello and colleagues figured there might be a cellular program that regulates lysosomal activity. Consistent with this idea, they found, using a Web-based tool for characterizing gene lists from high-throughput studies (g:Profiler; see Reimand et al., 2007), that genes encoding lysosomal proteins do tend to have coordinated expression. Furthermore, they identified a 10-base pair palindromic sequence located within 200 base pairs of the transcriptional start site in 68 of 96 known lysosomal genes, suggesting that the sequence is part of a promoter. The scientists put this motif (which they dubbed CLEAR, for Coordinated Lysosomal Expression And Regulation) through luciferase reporter assays and showed it could drive transcriptional activation.

Noting overlap between the CLEAR consensus sequence and the target site for four basic helix-loop-helix (bHLH) transcription factors, Ballabio’s team transfected HeLa cells with the four genes and found that overexpression of one, TFEB, resulted in elevated levels of lysosomal mRNAs. This was accompanied by corresponding increases in the activities of three lysosomal enzymes (β-glucosidase, cathepsin D, and β-glucoronidase).

Using chromatin immunoprecipitation analysis, the researchers confirmed that TFEB binds directly to CLEAR sites. They did microarray analysis of TFEB-overexpressing HeLa cells and found some 300 genes that appeared responsive to the factor. The vast majority of these genes were upregulated in response to TFEB, contained CLEAR elements in their promoters, and encoded proteins involved in lysosomal biogenesis and function. In this analysis, TFEB also appears to modulate the expression of several non-lysosomal genes involved with intracellular degradation, e.g., the autophagy factors Ras-related GTP binding C (RRAGC) and ultraviolet radiation resistance associated gene (UVRAG). Lysosomal degradation is the final step of the autophagy pathway, by which cells recycle damaged organelles and aggregated proteins.

Peering into TFEB-overexpressing HeLa cells using electron microscopy, the researchers found greater numbers of lysosomes in these stable transfectants, suggesting that TFEB promotes lysosome biogenesis. Using pulse-chase experiments, they determined that TFEB-overexpressing cells were able to degrade glycosaminoglycans faster than did mock-transfected cells. This heightened degradation capability could be relevant to neurodegenerative disease: a rat striatal cell model (HD43) with an inducible huntingtin transgene accumulated less huntingtin protein if transfected with TFEB, the researchers found.

“The potential for genetically inducing the lysosomal system to flush undesirable proteins more quickly from cells is certainly an exciting prospect,” wrote Ralph Nixon of New York University School of Medicine, who was not involved with the study, in an e-mail to ARF (see full comment below). “It highlights the need for this mobilized lysosomal system to function efficiently in the face of increasing challenges, such as cellular aging, gene mutations, and other factors that promote lysosomal impairment in Alzheimer disease, Parkinson disease, and other disorders.” Previous work has shown that boosting autophagy can prevent aggregation of mutant huntingtin and rescue HD-like behavioral symptoms in flies and mice (Ravikumar et al., 2004 and ARF related news story). Scientists have also identified a key autophagy regulator (beclin-1) that seems to help AD mice accumulate less Aβ (Pickford et al., 2008 and ARF related news story).

Nixon noted that the CLEAR network includes presenilin and nicastrin, two key components of the γ-secretase complex, which helps unleash the Aβ peptides in amyloid plaques. This “supports evidence that these proteins are enriched in lysosomal compartments and may well have roles in lysosomal degradative functions relevant to AD pathogenesis,” he wrote.—Esther Landhuis.

Reference:
Sardiello M, Palmieri M, di Ronza A, Medina DL, Valenza M, Gennarino VA, Di Malta C, Donaudy F, Embrione V, Polishchuk RS, Banfi S, Parenti G, Cattaneo E, Ballabio A. A Gene Network Regulating Lysosomal Biogenesis and Function. Science. 2009 Jun 25. Abstract