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Insulin-like growth factor-binding protein (IGFBP)-3 and IGFBP-5 share a common nuclear transport pathway in T47D
L J Schedlich1, T F Young, S M Firth
1Kolling Institute of Medical Research, University of Sydney, Royal North Shore Hospital, Sydney, New South Wales 2065, Australia. lyns@med.usyd.edu.au
Abstract:
Insulin-like growth factor-binding proteins (IGFBPs) play an integral role in modifying insulin-like growth factor actions in a wide variety of cell types. Recent evidence suggests that IGFBP-3 and IGFBP-5 also have effects on cell growth that are insulin-like growth factor-independent. In investigating possible mechanisms for this effect, the intracellular trafficking of IGFBP-3 and IGFBP-5, both of which contain sequences with the potential for nuclear localization, was studied in T47D cells. Nuclear uptake of fluorescently labeled IGFBP-3 and IGFBP-5 was observed in a proportion of T47D cells that appeared to be rapidly dividing. IGFBP-1 and IGFBP-2, which do not possess the putative domain for nuclear translocation, were not transported to the nuclei of T47D cells. When T47D cells were preincubated with excess unlabeled IGFBP-3, nuclear localization of labeled IGFBP-3 or IGFBP-5 was not detected, indicating that their nuclear translocation involves a common pathway. Inhibition of receptor-mediated endocytosis did not affect nuclear uptake of IGFBP-3, suggesting that it uses an alternative non-classical import pathway for transport across the plasma membrane. In addition, a variant form of IGFBP-3 with a mutation in the putative nuclear localization sequence was unable to translocate to the nuclei of T47D cells, suggesting that nuclear translocation of IGFBP-3 was dependent on these carboxyl-terminal basic residues.
Insights
Insulin-like growth factor-binding proteins (IGFBPs) can enter the nucleus independently of insulin-like growth factor signaling. This nuclear transport of IGFBP-3 and IGFBP-5 is crucial for their cell growth effects.
Area of Science:
- Cell Biology
- Molecular Endocrinology
- Cancer Research
Background:
- Insulin-like growth factor-binding proteins (IGFBPs) modulate insulin-like growth factor (IGF) actions.
- IGFBP-3 and IGFBP-5 exhibit IGF-independent effects on cell growth.
- The intracellular trafficking of IGFBPs is being investigated for novel mechanisms of action.
Purpose of the Study:
- To investigate the intracellular trafficking and nuclear localization of IGFBP-3 and IGFBP-5 in T47D cells.
- To determine if nuclear translocation of IGFBPs is dependent on IGF signaling.
- To elucidate the pathway involved in IGFBP nuclear import.
Main Methods:
- Utilized fluorescently labeled IGFBP-3 and IGFBP-5 for intracellular tracking in T47D cells.
- Compared nuclear uptake of IGFBPs with and without nuclear localization sequences (NLS).
- Investigated the role of receptor-mediated endocytosis and competitive inhibition using unlabeled IGFBP-3.
Main Results:
- Nuclear uptake of IGFBP-3 and IGFBP-5 was observed in a subset of rapidly dividing T47D cells.
- IGFBP-1 and IGFBP-2, lacking a putative NLS, did not show nuclear transport.
- Nuclear translocation of IGFBP-3 and IGFBP-5 involved a common pathway, dependent on the NLS of IGFBP-3 and independent of endocytosis.
Conclusions:
- IGFBP-3 and IGFBP-5 can translocate to the nucleus in T47D cells.
- Nuclear import of IGFBP-3 and IGFBP-5 is mediated by a specific pathway involving their NLS.
- These findings suggest novel, IGF-independent roles for IGFBPs in nuclear functions and cell growth regulation.