Résumé

It has long been known that liver lysosomes contain an endoglycosidase activity able to degrade the high molecular mass glycosaminoglycan hyaluronic acid (HA). The identification and cloning of a hyaluronidase with an acidic pH optimum, Hyal-1, suggested it might be responsible for this activity. However, we previously reported that this hydrolase could only be detected in pre-lysosomal compartments of the mouse liver using a zymography technique that allows the detection of Hyal-1 activity after SDS-PAGE ("renatured protein zymography"). Present work reveals that the activity highlighted by this technique belongs to a precursor form of Hyal-1 and that the lysosomal HA endoglycosidase activity of the mouse liver is accounted for by a proteolytically processed form of Hyal-1 that can only be detected using "native protein zymography". Indeed, the distribution of this form follows the distribution of β-galactosidase, a well-established lysosomal marker, after fractionation of the mouse liver in a linear sucrose density gradient. In addition, both activities shift toward the lower density region of the gradient when a specific decrease of the lysosomal density is induced by Triton WR-1339 injection. The fact that only native protein zymography but not renatured protein zymography is able to detect Hyal-1 activity in lysosomes points to a non-covalent association of Hyal-1 proteolytic fragments or the existence of closely linked partners supporting Hyal-1 enzymatic activity. The knockdown of Hyal-1 results in an 80% decrease of total acid hyaluronidase activity in the mouse liver, confirming that Hyal-1 is a key actor of HA catabolism in this organ.

langue originaleAnglais
Pages (de - à)1155-60
Nombre de pages6
journalBiochemical and Biophysical Research Communications
Volume446
Numéro de publication4
Les DOIs
étatPublié - 2014

Empreinte digitale

Lysosomes
Liver
Hyaluronic Acid
Hyaluronoglucosaminidase
Glycoside Hydrolases
Proteins
Galactosidases
Cloning
Molecular mass
Hydrolases
Fractionation
Glycosaminoglycans
Sucrose
Organism Cloning
Polyacrylamide Gel Electrophoresis
Injections
Acids

Citer ceci

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title = "Mouse liver lysosomes contain enzymatically active processed forms of Hyal-1",
abstract = "It has long been known that liver lysosomes contain an endoglycosidase activity able to degrade the high molecular mass glycosaminoglycan hyaluronic acid (HA). The identification and cloning of a hyaluronidase with an acidic pH optimum, Hyal-1, suggested it might be responsible for this activity. However, we previously reported that this hydrolase could only be detected in pre-lysosomal compartments of the mouse liver using a zymography technique that allows the detection of Hyal-1 activity after SDS-PAGE ({"}renatured protein zymography{"}). Present work reveals that the activity highlighted by this technique belongs to a precursor form of Hyal-1 and that the lysosomal HA endoglycosidase activity of the mouse liver is accounted for by a proteolytically processed form of Hyal-1 that can only be detected using {"}native protein zymography{"}. Indeed, the distribution of this form follows the distribution of β-galactosidase, a well-established lysosomal marker, after fractionation of the mouse liver in a linear sucrose density gradient. In addition, both activities shift toward the lower density region of the gradient when a specific decrease of the lysosomal density is induced by Triton WR-1339 injection. The fact that only native protein zymography but not renatured protein zymography is able to detect Hyal-1 activity in lysosomes points to a non-covalent association of Hyal-1 proteolytic fragments or the existence of closely linked partners supporting Hyal-1 enzymatic activity. The knockdown of Hyal-1 results in an 80{\%} decrease of total acid hyaluronidase activity in the mouse liver, confirming that Hyal-1 is a key actor of HA catabolism in this organ.",
keywords = "Animals, Gene Knockdown Techniques, Hyaluronic Acid, Hyaluronoglucosaminidase, Liver, Lysosomes, Mice, Mice, Inbred C57BL",
author = "Marielle Boonen and Emeline Puissant and Florentine Gilis and Bruno Flamion and Michel Jadot",
note = "Copyright {\circledC} 2014 Elsevier Inc. All rights reserved.",
year = "2014",
doi = "10.1016/j.bbrc.2014.03.070",
language = "English",
volume = "446",
pages = "1155--60",
journal = "Biochemical and Biophysical Research Communications",
issn = "0006-291X",
publisher = "Academic Press Inc.",
number = "4",

}

TY - JOUR

T1 - Mouse liver lysosomes contain enzymatically active processed forms of Hyal-1

AU - Boonen, Marielle

AU - Puissant, Emeline

AU - Gilis, Florentine

AU - Flamion, Bruno

AU - Jadot, Michel

N1 - Copyright © 2014 Elsevier Inc. All rights reserved.

PY - 2014

Y1 - 2014

N2 - It has long been known that liver lysosomes contain an endoglycosidase activity able to degrade the high molecular mass glycosaminoglycan hyaluronic acid (HA). The identification and cloning of a hyaluronidase with an acidic pH optimum, Hyal-1, suggested it might be responsible for this activity. However, we previously reported that this hydrolase could only be detected in pre-lysosomal compartments of the mouse liver using a zymography technique that allows the detection of Hyal-1 activity after SDS-PAGE ("renatured protein zymography"). Present work reveals that the activity highlighted by this technique belongs to a precursor form of Hyal-1 and that the lysosomal HA endoglycosidase activity of the mouse liver is accounted for by a proteolytically processed form of Hyal-1 that can only be detected using "native protein zymography". Indeed, the distribution of this form follows the distribution of β-galactosidase, a well-established lysosomal marker, after fractionation of the mouse liver in a linear sucrose density gradient. In addition, both activities shift toward the lower density region of the gradient when a specific decrease of the lysosomal density is induced by Triton WR-1339 injection. The fact that only native protein zymography but not renatured protein zymography is able to detect Hyal-1 activity in lysosomes points to a non-covalent association of Hyal-1 proteolytic fragments or the existence of closely linked partners supporting Hyal-1 enzymatic activity. The knockdown of Hyal-1 results in an 80% decrease of total acid hyaluronidase activity in the mouse liver, confirming that Hyal-1 is a key actor of HA catabolism in this organ.

AB - It has long been known that liver lysosomes contain an endoglycosidase activity able to degrade the high molecular mass glycosaminoglycan hyaluronic acid (HA). The identification and cloning of a hyaluronidase with an acidic pH optimum, Hyal-1, suggested it might be responsible for this activity. However, we previously reported that this hydrolase could only be detected in pre-lysosomal compartments of the mouse liver using a zymography technique that allows the detection of Hyal-1 activity after SDS-PAGE ("renatured protein zymography"). Present work reveals that the activity highlighted by this technique belongs to a precursor form of Hyal-1 and that the lysosomal HA endoglycosidase activity of the mouse liver is accounted for by a proteolytically processed form of Hyal-1 that can only be detected using "native protein zymography". Indeed, the distribution of this form follows the distribution of β-galactosidase, a well-established lysosomal marker, after fractionation of the mouse liver in a linear sucrose density gradient. In addition, both activities shift toward the lower density region of the gradient when a specific decrease of the lysosomal density is induced by Triton WR-1339 injection. The fact that only native protein zymography but not renatured protein zymography is able to detect Hyal-1 activity in lysosomes points to a non-covalent association of Hyal-1 proteolytic fragments or the existence of closely linked partners supporting Hyal-1 enzymatic activity. The knockdown of Hyal-1 results in an 80% decrease of total acid hyaluronidase activity in the mouse liver, confirming that Hyal-1 is a key actor of HA catabolism in this organ.

KW - Animals

KW - Gene Knockdown Techniques

KW - Hyaluronic Acid

KW - Hyaluronoglucosaminidase

KW - Liver

KW - Lysosomes

KW - Mice

KW - Mice, Inbred C57BL

U2 - 10.1016/j.bbrc.2014.03.070

DO - 10.1016/j.bbrc.2014.03.070

M3 - Article

C2 - 24667601

VL - 446

SP - 1155

EP - 1160

JO - Biochemical and Biophysical Research Communications

JF - Biochemical and Biophysical Research Communications

SN - 0006-291X

IS - 4

ER -