Project: Research project

Project Details


DESCRIPTION (Verbatim from the applicant's abstract): The investigator will
study membrane lipid transport and sphingolipid/cholesterol interactions in
sphingolipid storage disease (SLSD) fibroblasts. Recently he found that several
fluorescent SL analogs were internalized from the plasma membrane (PM)
predominantly to the Golgi complex of normal cells, while in 10 different SLSD
cell types, these lipids accumulated in endosomes and lysosomes. This
accumulation was in general unrelated to the extent of SL analog degradation in
the different cell types. He also showed that cholesterol bomeostasis is
perturbed in multiple SLSDs secondary to SL accumulation and that mistargeting
of SL analogs was regulated by cholesterol. Based on these results, they
hypothesize that endogenous lipids which accumulate in SLSD cells due to
primary defects in lipid catabolism result in an altered intracellular
distribution of cholesterol, and that this alteration in membrane composition
then results in defective sorting and transport of SLs. Four broad projects
pertaining to this hypothesis pursued. They will (i) examine the
itinerary of fluorescent internalized recycled from the PM to various
intracellular compartments over time: Dominant negative constructs of several
different Rab proteins will be used to help define the compartment(s) where
defective SL sorting occurs; (ii) study the intracellular transport of
endogenous (e.g., using fluorescent SL binding toxins, anti-SL antibodies, or
resialylation of SLs) to confirm their preliminary observations that the
perturbation in PM to Golgi traffic seen in SLSD fibroblasts with fluorescent
SLs is mirrored by endogenous SLs; (iii) evaluate potential mechanisms
responsible for the perturbation of cholesterol homeostasis in SLSD cells. In
particular, they will determine whether the ability of exogenous SLs to perturb
cholesterol homeostasis in cells is related to the strength of specific
SL/cholesterol interactions in vitro. They will also evaluate the potential
roles of several sterol-sensing proteins in modulating cholesterol homeostasis
in SLSD cells; and (iv) evaluate mechanistic hypotheses modulating Sl transport
targeting in SLSD cells. The role of "membrane fluidity" will be studied by
modifying endogenous membrane lipids and by systematically varying the chain
length and hydrophobicity of the analogs used to observe SL sorting in SLSD
cells. They will also examine the function of the sterol transport protein, NPC
1, suggested by their preliminary data to play a critical role in SL targeting.
These studies will test a novel concept concerning the involvement of
SL/cholesterol interactions in the regulation of lipid trafficking in normal
and SLSD cells.
Effective start/end date4/1/0011/30/12


  • National Institute of General Medical Sciences: $296,186.00
  • National Institute of General Medical Sciences: $299,178.00


  • Medicine(all)
  • Biochemistry, Genetics and Molecular Biology(all)


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