Biochemical characterization and quantitative gene expression analysis of the multi-stress inducible metallothionein from Tetrahymena thermophila.
A cadmium-binding protein with biochemical features of a metallothionein (MT) has been isolated and purified to homogeneity from the ciliate Tetrahymena thermophila. N-terminal sequencing revealed the posttranslational cleavage of the first two amino acids and, in general, a high degree of identity with known MTs from other ciliates. Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) analysis of the apothionein revealed a molecular mass (16,763 Da) higher to those of mammals and of other protozoa. Finally, quantitative real-time PCR has been used to investigate the susceptibility of this ciliate MT to gene activation in response to heavy metals and to other stressors. Our data indicate that while zinc is not effective at all and cadmium is the best inducer, other stress factors, such as mercury, copper, heat and hydrogen peroxide, also activated gene transcription. As in vertebrate cells, interleukin-6 (IL-6) that stimulates ciliate growth, was able to enhance MT gene synthesis. This complex of data seems to indicate a general role of this protein in stress response.
Department of Science and Advanced Technology, University of Piemonte Orientale "Amedeo Avogadro", Corso Borsalino, Alessandria, Italy. fdondero@unipmn.it
Francesco
F
Dondero
Maria
M
Cavaletto
Anna Rita
AR
Ghezzi
Antonietta
A
La Terza
Mohamed
M
Banni
Aldo
A
Viarengo
D000818:Animals; D008668:Metallothionein / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D016133:Polymerase Chain Reaction; D011499:Protein Processing, Post-Translational; D012044:Regression Analysis; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D016808:Tetrahymena thermophila / Q000235:genetics* / Q000502:physiology
D008668:Metallothionein