N-terminal cleavage of proTGFalpha occurs at the cell surface by a TACE-independent activity.
ProTGFalpha (transforming growth factor alpha precursor) maturation and conversion into soluble TGFalpha is a complex process that involves three proteolytic steps. One, that occurs co-translationally, eliminates the signal sequence. Another, occurring at the juxta-membrane domain, solubilizes TGFalpha. A third cleavage removes the N-terminal extension of proTGFalpha. This latter step has been poorly studied, mainly because of the rapid kinetics of this cleavage. In the present study, we have designed a strategy to analyse several aspects regarding this N-terminal cleavage. In vivo treatment with the hydroxamate-based metalloprotease inhibitors BB3103 or TAPI-2 (tumour necrosis factor-alpha protease inhibitor 2) reversibly induced accumulation of forms of proTGFalpha that included the N-terminal extension. N-terminal shedding was rapid, and occurred at the cell surface. However, the machinery responsible for the N-terminal cleavage was inactive in other cellular sites, such as the endoplasmic reticulum. Experiments of proTGFalpha expression and maturation in cells deficient in TACE (tumour-necrosis-factor-alpha-converting enzyme) activity indicated that this protease was dispensable for N-terminal processing of proTGFalpha in vivo, but was required for regulated cleavage at the C-terminus. These findings indicate that TACE is not involved in N-terminal processing of proTGFalpha, and suggest differences in the machineries that control the cleavage at both ends of TGFalpha within its precursor.
Juanes
Pedro P
PP
Instituto de Microbiología Bioquímica and Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Campus Miguel de Unamuno, 37007-Salamanca, Spain.
Ferreira
Laura
L
Montero
Juan Carlos
JC
Arribas
Joaquín
J
Pandiella
Atanasio
A
D051722:ADAM Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000072198:ADAM17 Protein; D053829:Amyloid Precursor Protein Secretases; D000818:Animals; D016282:Aspartic Acid Endopeptidases; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism*; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D010450:Endopeptidases / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D011498:Protein Precursors / Q000737:chemistry / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D016211:Transforming Growth Factor alpha / Q000737:chemistry / Q000378:metabolism*
D006877:Hydroxamic Acids; D011480:Protease Inhibitors; D011498:Protein Precursors; D016211:Transforming Growth Factor alpha; C052268:protransforming growth factor alpha; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human; D051722:ADAM Proteins; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human