The Gene for a Novel Member of the Whey Acidic Protein Family Encodes Three Four-disulfide Core Domains and Is Asynchronously Expressed during Lactation

Abstract

Secretion of whey acidic protein (WAP) in milk throughout lactation has previously been reported for a limited number of species, including the mouse, rat, rabbit, camel, and pig. We report here the isolation of WAP from the milk of a marsupial, the tammar wallaby (Macropus eugenii). Tammar WAP (tWAP) was isolated by reverse-phase HPLC and migrates in SDS-polyacrylamide gel electrophoresis at 29.9 kDa. tWAP is the major whey protein, but in contrast to eutherians, secretion is asynchronous and occurs only from approximately days 130 through 240 of lactation. The full-length cDNA codes for a mature protein of 191 amino acids, which is comprised of three four-disulfide core domains, contrasting with the two four-disulfide core domain arrangement in all other known WAPs. A three-dimensional model for tWAP has been constructed and suggests that the three domains have little interaction and could function independently. Analysis of the amino acid sequence suggests the protein belongs to a family of protease inhibitors; however, the predicted active site of these domains is dissimilar to the confirmed active site for known protease inhibitors. This suggests that any putative protease ligand may be unique to either the mammary gland, milk, or gut of the pouch young. Examination of the endocrine regulation of the tWAP gene showed consistently that the gene is prolactin-responsive but that the endocrine requirements for induction and maintenance of tWAP gene expression are different during lactation.

Inhibitory Function of Whey Acidic Protein in the Cell-Cycle Progression of Mouse Mammary Epithelial Cells (EpH4/K6 Cells)

“…For example, it has been suggested that WAP may be a source of cysteine for hair growth of pouch young because WAP contains a large number of cysteine residues [9]. It has been also suggested that WAP may contribute, via milk, to the development of hair in rodents, which are born with minimal fur coverage [7]. On the other hand, Nicholas et al [8] has suggested that WAP may influence the rate of development of the gut of the young via milk.…”

“…The forced expression of exogenous WAP significantly impaired the proliferation of EpH4 cells, whereas it did not affect that of . Recently, it has also isolated from the milk of pigs [5,6], and three marsupial species, tamer wallaby [7], red kangaroo [8], and brushtail possum [9]. Expression of the WAP gene is regulated by lactogenic hormones, and the level of mo use WAP m RN A in the m am m ary gla nds incr eases t hous ands -fo ld between non-lactation and mid-lactation [10].…”

Identification, Evolution, and Regulation of Expression of Guinea Pig Trappin with an Unusually Long Transglutaminase Substrate Domain

“…This four-disulfide core structure was first described in whey acidic protein in mouse milk and thus was named the "WAP motif" (7). Members of the family of WAP motif proteins contain one to three WAP motifs, many of which have proteinase inhibitor activity and/or antibacterial activity (8,9). Trappins are unique in having TGS domains in their N termini among diverse family members of WAP motif-containing proteins that include toxins, pollen allergens, serine proteinase inhibitors, growth inhibitors, calcium transport inhibitors, and others whose functions have not yet been determined (10 -17).…”

Analysis of the Promoter of Mutated Human Whey Acidic Protein (WAP) Gene

“…hey acidic protein (WAP) has been identified in the milk of a wide range of laboratory animals rodents [1,2], as well as in camels [3], rabbits [4], pigs [5], and three marsupial species, the tammar wallaby [6], brushtail possum [7], the red kangaroo [8]. In mice and rabbits, the expression of the WAP gene occurs in mammary epithelial cells during late pregnancy and throughout lactation, and is regulated by a variety of lactogenic hormones, including insulin, glucocorticoid, and prolactin [9][10][11][12][13][14].…”