The Fungus Tremella mesenterica Encodes the Longest Metallothionein Currently Known: Gene, Protein and Metal Binding Characterization

Abstract

Fungal Cu-thioneins, and among them, the paradigmatic Neurospora crassa metallothionein (MT) (26 residues), were once considered as the shortest MTs -the ubiquitous, versatile metal-binding proteins- among all organisms, and thus representatives of their primeval forms. Nowadays, fungal MTs of diverse lengths and sequence features are known, following the huge heterogeneity of the Kingdom of Fungi. At the opposite end of N. crassa MT, the recently reported Cryptococcus neoformans CnMT1 and CnMT2 (122 and 186 aa) constitute the longest reported fungal MTs, having been identified as virulence factors of this pathogen. CnMTs are high-capacity Cu-thioneins that appear to be built by tandem amplification of a basic unit, a 7-Cys segment homologous to N. crassa MT. Here, we report the in silico, in vivo and in vitro study of a still longer fungal MT, belonging to Tremella mesenterica (TmMT), a saprophytic ascomycete. The TmMT gene has 10 exons, and it yields a 779-bp mature transcript that encodes a 257 residue-long protein. This MT is also built by repeated fragments, but of variable number of Cys: six units of the 7-Cys building blocks-CXCX3CSCPPGXCXCAXCP-, two fragments of six Cys, plus three Cys at the N-terminus. TmMT metal binding abilities have been analyzed through the spectrophotometric and spectrometric characterization of its recombinant Zn-, Cd- and Cu-complexes. Results allow it to be unambiguous classified as a Cu-thionein, also of extraordinary coordinating capacity. According to this feature, when the TmMT cDNA is expressed in MT-devoid yeast cells, it is capable of restoring a high Cu tolerance level. Since it is not obvious that T. mesenterica shares the same physiological needs for a high capacity Cu-binding protein with C. neoformans, the existence of this peculiar MT might be better explained on the basis of a possible role in Cu-handling for the Cu-enzymes responsible in lignin degradation pathways.

Cadmium exposure during pregnancy and lactation: materno-fetal and newborn repercussions of Cd(ii), and Cd–metallothionein complexes

“…MTs are a superfamily of ubiquitous, small to medium-sized (o30 kDa) 14,15 cysteine-rich (15-30%) proteins with the ability to coordinate metal ions through metal-thiolate bonds and have been identified in all eukaryotes and some prokaryotes thus far analysed. [16][17][18] The variety in the metal ion coordination that MTs show, reveal particularities in the three-dimensional metal-MT complexes.…”

Cloning and functional characterization of a novel metallothionein gene in Antarctic sea‐ice yeast (Rhodotorula mucilaginosa)

“…First, the RmMT gene encoded 106 amino acids, longer than general MT (ca. 60 aa) , but shorter than TmMT (257 aa) of Tremella mesenterica , CnMT1 (122 aa) and CnMT2 (183 aa) of Cryptoccocus neoformans . Second, 16 cysteine residues were scattered throughout the whole sequence and its content in RmMT was about 15%, much lower than typical MTs (ca.…”

Cadmium Uptake, MT Gene Activation and Structure of Large-Sized Multi-Domain Metallothioneins in the Terrestrial Door Snail Alinda biplicata (Gastropoda, Clausiliidae)

“…Intron loss can be a sign for pseudogenization [46], meaning that the respective gene is not transcribed into mRNA any more, which normally serves as a template for translation into the respective protein. [47]; H.p., Helix pomatia [13]; B.g., Biomphalaria glabrata [48]; A.b., Alinda biplicata (this study); T.m., Tremella mesenterica [49]; C.v., Crassostrea virginica [50]; T.p., Tetrahymena pyriformis [51].…”

“…All these features argue against the pseudogenization of the two intronless MT genes. Overall, the internal architecture of MT genes can be very variable ( Figure 3), and three different kinds of MT gene structures can be distinguished: (i) a typical exon/intron structure as known for many other genes, too [13,[47][48][49][50]; (ii) a deviated exon/intron structure with a large exon consisting of repeat sequences that sometimes represent multiple domains [47]; and (iii) real intronless genes where introns are not present anymore [51] (this study). In addition, the length of the introns can vary to a great extent, mostly in contrast to the rather uniform size of exons ( Figure 3).…”