In mammals a single receptor protein binds both insulin-like growth factor II (IGF-II) and mannose 6-phosphate (Man 6-P) containing ligands, most notably lysosomal enzymes. However, in chick embryo fibroblasts IGF-II binds predominantly to a type 1 IGF receptor, and no IGF-II/Man 6-P receptor has been identified in this species. In order to determine if chickens possess an IGF-II/Man 6-P receptor, an affinity resin (pentamannosyl 6-phosphate (PMP) Sepharose) was used to purify receptors from chicken membrane extracts by their ability to bind mannose 6-phosphate. Then 125I-IGF-II was used to evaluate their ability to bind IGF-II. These experiments demonstrate that nonmammalian Man 6-P receptors lack the ability to bind IGF-II, suggesting that the ability to bind IGF-II has been gained recently in evolution by the mammalian Man 6-P receptor.
The second area of study involves the serum form of the IGF-II/Man 6-P receptor. This receptor had been detected in the serum of a number of mammalian species, yet its structure, function, regulation, and origin were unknown. Initial studies, done with Dr. R. G. MacDonald, showed that the serum receptor is truncated such that the C-terminal cytoplasmic domain of the cellular receptor is removed. These studies also demonstrate a regulation of serum receptor levels with age, similar to that seen for the cellular receptor, and that the serum form of the receptor existed in several forms which appeared intact under nonreducing conditions, but as multiple proteolytic products upon reduction. Finally, these studies demonstrated that both the cellular and serum IGF-II/Man 6-P receptors are capable of binding IGF-II and Man 6-P simultaneously.
In studies on the serum form of the IGF-II/Man 6-P receptor that I have conducted independently, the regulation of the serum IGF-II/Man 6-P and transferrin receptors by insulin has been demonstrated. In these studies, insulin injected into rats subcutaneously resulted in a time and dose dependent increase in serum receptor levels. Finally, to investigate the relationship of the serum IGF- II/Man 6-P receptor to the cellular form of the receptor, pulse chase experiments were performed. These experiments demonstrate that the soluble (serum form released into the medium) receptor is a major degradation product of the cellular receptor. Furthermore, the lack of detectable amounts of the lower Mr soluble receptor intracellularly and the parallel relationship of cell surface and soluble receptor suggest that the proteolysis is occurring from the cell surface. Finally, a number of experiments suggest that the degradation rate depends upon the conformation state of the receptor: binding of IGF-II or Man 6-P makes the receptor more susceptible to proteolysis while the presence of lysosomal enzymes prevents receptor proteolysis.
In summary, the serum form of the IGF-II receptor is a proteolytic product of the cellular form of the receptor. The rate of release depends upon the number of receptors at the cell surface and the binding state of the receptor. In circulation, the receptor retains the ability to bind both types of ligands, it thus may serve as an IGF binding protein and/or a lysosomal enzyme binding protein. These results suggest a model whereby the cellular receptor is proteolytically cleaved by a plasma membrane protease to produce a short membrane anchored fragment and the serum receptor. In vivo this pathway serves as the major degradative pathway of the IGF-II/Man 6-P receptor, with the serum form being cleared from circulation by further degradation and reuptake.
| Identifer | oai:union.ndltd.org:umassmed.edu/oai:escholarship.umassmed.edu:gsbs_diss-1044 |
| Date | 01 October 1990 |
| Creators | Clairmont, Kevin B. |
| Publisher | eScholarship@UMMS |
| Source Sets | University of Massachusetts Medical School |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | GSBS Dissertations and Theses |
| Rights | Copyright is held by the author, with all rights reserved., select |
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