It has been suggested that intracellular Hyal-1 (hyaluronidase-1), which is considered a lysosomal enzyme, originates via endocytosis of the serum enzyme. To test this proposal we have investigated the uptake and intracellular distribution of rhHyal-1 (recombinant human Hyal-1) by mouse liver, making use of centrifugation methods. Experiments were performed on wild-type mice injected with 125I-labelled rhHyal-1 and on Hyal-1−/− mice injected with the unlabelled enzyme, which were killed at various times after injection. Activity of the unlabelled enzyme was determined by zymography. Intracellular distribution of Hyal-1 was investigated by differential and isopycnic centrifugation. The results of the study indicated that rhHyal-1 is endocytosed by the liver, mainly by sinusoidal cells, and follows the intracellular pathway described for many endocytosed proteins that are eventually located in lysosomes. However, Hyal-1 endocytosis has some particular features. First, endocytosed rhHyal-1 is quickly degraded. Secondly, its distribution, as analysed by differential centrifugation, differs from the distribution of β-galactosidase, taken as the reference lysosomal enzyme. Further analysis by isopycnic centrifugation in a sucrose gradient shows endocytosed rhHyal-1 behaves like β-galactosidase shortly after injection. However the Hyal-1 distribution is markedly less affected than β-galactosidase, following a prior injection of Triton WR-1339, which is a specific density perturbant of lysosomes. The behaviour in centrifugation of endogenous liver Hyal-1, identified by hyaluronan zymography, exhibits some similarity with the behaviour of the endocytosed enzyme, suggesting that it could originate from endocytosis of the serum enzyme. Overall, these results can be explained by supposing that active endocytosed Hyal-1 is mainly present in early lysosomes. Although its degradation half-time is short, Hyal-1 could exert its activity due to a constant supply of active molecules from the blood.
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Research Article|
August 13 2010
Endocytosis of hyaluronidase-1 by the liver
Marie-Christine Gasingirwa;
Marie-Christine Gasingirwa
1
*Laboratory of Physiological Chemistry, URPhyM, University of Namur, Namur, Belgium
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Jacqueline Thirion;
Jacqueline Thirion
*Laboratory of Physiological Chemistry, URPhyM, University of Namur, Namur, Belgium
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Jeannine Mertens-Strijthagen;
Jeannine Mertens-Strijthagen
†Laboratory of Physiology and Pharmacology, URPhyM, University of Namur, Namur, Belgium
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Simone Wattiaux-De Coninck;
Simone Wattiaux-De Coninck
*Laboratory of Physiological Chemistry, URPhyM, University of Namur, Namur, Belgium
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Bruno Flamion;
Bruno Flamion
†Laboratory of Physiology and Pharmacology, URPhyM, University of Namur, Namur, Belgium
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Robert Wattiaux;
Robert Wattiaux
2
*Laboratory of Physiological Chemistry, URPhyM, University of Namur, Namur, Belgium
2To whom correspondence should be addressed (email robert.wattiaux@hotmail.com).
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Michel Jadot
Michel Jadot
*Laboratory of Physiological Chemistry, URPhyM, University of Namur, Namur, Belgium
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Publisher: Portland Press Ltd
Received:
May 12 2010
Revision Received:
June 16 2010
Accepted:
June 17 2010
Accepted Manuscript online:
June 17 2010
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2010 Biochemical Society
2010
Biochem J (2010) 430 (2): 305–313.
Article history
Received:
May 12 2010
Revision Received:
June 16 2010
Accepted:
June 17 2010
Accepted Manuscript online:
June 17 2010
Citation
Marie-Christine Gasingirwa, Jacqueline Thirion, Jeannine Mertens-Strijthagen, Simone Wattiaux-De Coninck, Bruno Flamion, Robert Wattiaux, Michel Jadot; Endocytosis of hyaluronidase-1 by the liver. Biochem J 1 September 2010; 430 (2): 305–313. doi: https://doi.org/10.1042/BJ20100711
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