SUMMARY PARAGRAPH for MAC1
MAC1 encodes a transcription factor ("Metal-binding activator") that regulates the expression of genes involved in copper homeostasis (1). In response to low copper levels, Mac1p induces expression of the copper transporters Ctr1p (2, 3, 4, 5, 6) and Ctr3p (3, 5, 6), the (known or putative) cell-surface metal reductases Fre1p (1, 2, 3, 4, 5, 6, 7, 8, 9, 10) and Fre7p (6, 9, 10) and two uncharacterized ORF's, YFR055W and YJL217W (6). FRE1 is expressed early and FRE2 is expressed late during copper depletion; in Mac1p-depleted cells, FRE1 expression persists for longer and FRE2 expression begins earlier (8). Mac1p is also responsible for the H2O2-induced transcription of CTT1 (1). Copper inhibits the activity of Mac1p (2, 7, 8).
Cells lacking active Mac1p are viable but copper-deficient, respiratory deficient, and sensitive to heat, hydrogen peroxide, cadmium, zinc, and lead; normal growth and respiration can be restored by supplementing the growth medium with copper or iron (1). Mac1p-depleted cells are also sensitive to oxytetracycline and doxycycline (11) and resistant to the anti-cancer drug cisplatin (12). Since iron uptake depends heavily on copper uptake, cells lacking active Mac1p also exhibit a secondary iron deficiency (7, 13). A hypermorphic mutation, MAC1up1, is resistant to inhibition by copper (7) and causes increased copper uptake and copper hypersensitivity (1, 5, 14).
Mac1p binds a copper-response element ("CuRE"), TTTGC(T/G)C(A/G), in the promoters of target genes (3, 4, 9, 10). Two CuREs are required for induction by Mac1p, and the spacing between them is important (9). Mac1p binds single CuREs as a monomer in vitro, but binds synergistically to promoters with two or more CuREs in vivo (15, 16). CuRE binding requires phosphorylation of Mac1p and is disrupted by Cu(I) or Ag(I). Mac1p also requires copper in order to bind CuREs, indicating that Mac1p detects both a minimum and a maximum allowable copper concentration (17); therefore, Mac1p is often referred to as the "nutritional" copper sensor, in contrast to the "toxic" copper sensor Cup2p (also known as Ace1p), which is activated when copper levels become excessive (5).
Mac1p has two nuclear localization signals, and the protein is found in the nucleus in both copper-starved and copper-replete cells (1, 18, 19). The N-terminal DNA-binding domain recognizes CuREs and binds two Zn(II) ions, which are required for DNA-binding activity (15, 16, 18, 19). Mac1p also has two cysteine-rich domains, referred to as Rep I and Rep II, that bind a total of eight copper ions (2, 18, 19). (Ultraviolet irradiation causes Mac1p to emit orange light, indicating that these are Cu(I) ions (18).) The constitutive MAC1up1 mutation lies in Rep I (2, 19) and the mutant protein binds only four or five copper ions (18); mutation of other residues in Rep I, but not Rep II, also leads to constitutive Mac1p activity (19, 20). Rep I binds four Cu(I) ions in a trigonal configuration similar to one found in Cup2p (21). Rep II is a copper-dependent transactivation domain (22). In the absence of copper, a C-terminal D helix mediates dimerization with other Mac1p molecules to form ternary activation complexes on the promoters of target genes (19). Nutritional copper levels stabilize Mac1p but cause an intramolecular reaction in which the D helix binds the DNA-binding domain and prevents binding to CuREs, abrogating gene activation (18, 19). Mac1up1p is more stable than the wild-type protein during copper depletion, but is not stabilized by copper (18). Mac1p is degraded at high copper concentrations, but MAC1up1p is resistant (14).
Mac1p also indirectly mediates the degradation of Ctr1p in high copper concentrations; the DNA-binding domain of Mac1p is required, and, as stated above, Mac1p is not found outside the nucleus, suggesting that Mac1p activates the transcription of genes required for Ctr1p degradation (23).
Last updated: 2005-08-24