Hutchinson-Gilford Progeria Syndrome (HGPS) is a premature aging disorder that is lethal by age 16. It is caused by a defect in a nuclear envelope protein called Lamin A. Lamins are structural proteins that line the inner surface of the nuclear envelope. Humans have two types of lamins: A-types (Lamin A and C) and B-types (Lamin B1 and B2). All lamins, except Lamin C, are processed for membrane anchorage by the addition of a hydrophobic prenyl (lipid) group at the C-terminus. But for unknown reasons, Lamin A is further processed to remove the anchor. Thus, Lamin A is transiently rather than stably prenylated. Mutations that cause HGPS delete a recognition site in the Lamin A protein required for removal of the anchor. This defective Lamin A protein is called progerin, and is stably rather than transiently prenylated, resulting in organism-wide rapid aging by unknown mechanisms. The enzyme thought to be responsible for removing the prenyl anchor from Lamin A is called Zmpste24, which is a zinc-dependant prenyl protease. The experiments described in this proposal will establish the fruit fly Drosophila as a model to study human Lamin A processing. It is possible to incorporate human genes into the fly genome (so-called transgenic flies). When transgenically expressed in flies, normal human Lamin A is stably, rather than transiently, anchored in the fly nuclear envelope, with no apparent ill effects. This is in contrast to the human situation, in which stable membrane anchorage of Lamin A leads to disease, and suggests that no conserved prenyl proteases in flies are removing the anchor from human Lamin A. Thus the Drosophila nuclear envelope can serve as an ideal environment in which to confirm the interaction between human Lamin A and the protease Zmpste24, which is a critical step that goes awry in HGPS.
In Specific Aim 1, normal or processing-defective human Lamin A proteins (including progerin) will be transgenically expressed in fruit flies together with human Zmpste24, and the localization patterns of Lamin A examined with immunofluorescence microscopy. Drosophila larvae possess huge, easily manipulated nuclei in their salivary glands, which permit detailed dissection of nuclear protein dynamics. Since human Lamin A is confined to the nuclear envelope in flies, co-expression with Zmpste24 should release membrane anchorage and cause re-localization into the nuclear interior, thus demonstrating a direct interaction between these two proteins.
Specific Aim 2 comprises a genetic and molecular dissection of the Drosophila prenyl protease functions, firstly to confirm they do not interfere with the human processing assay from Aim 1, and secondly to establish the degree to which lamin processing is conserved in flies. Results from this work will support the use of Drosophila as a non-human in vivo environment in which to study human lamin biology, and will reveal underlying generalities about the nuclear envelope and its role in aging.
Hutchinson-Gilford Progeria Syndrome is a devastating and extremely rare premature aging disorder that kills children before the age of 16. The research described in this proposal will contribute to world-wide efforts to find a cure or at least a palliative. But it is also very probable that unraveling the biology of this unusual disease will go some way to explaining the root causes of aging in general, a genetic disease which affects us all.
| Adolphsen, Katie; Amell, Amanda; Havko, Nathan et al. (2012) Type-I prenyl protease function is required in the male germline of Drosophila melanogaster. G3 (Bethesda) 2:629-42 |
