A marsupial β-lactoglobulin gene: characterization and prolactin-dependent expression

Abstract

Analysis of the tammar wallaby beta-lactoglobulin cDNA and inferred amino acid sequences reveal extensive sequence divergence from the eutherian beta-lactoglobulins. Conserved residues include the cysteines and a number of individual amino acids involved in structure and ligand-binding. The only region of extended similarity is a heptapeptide sequence which may impart specificity to its interaction with a receptor protein. Northern analysis of total mammary RNA revealed two transcripts which result from differential utilization of polyadenylation signals. The concentration of beta-lactoglobulin mRNA increased in late lactation and correlates with increases in milk production and levels of milk fat. Quantification of beta-lactoglobulin mRNA levels in hormone-stimulated mammary gland explants from tammars in late pregnancy suggests that maximal induction of the gene is dependent on prolactin alone and that expression is not modulated by other hormones known to play a role in the initiation of lactation in eutherians.

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

“…No tWAP mRNA was detected in explants cultured in media with either IF, P, or IFP. Expression of the ␤-LG gene was induced with P alone in mammary explants from tammars at day 24 of gestation and in all analyses was not detected after 4 days of culture in IF, confirming previous studies (22).…”

“…The tWAP and tammar ␤-lactoglobulin (␤-LG) cDNAs were hybridized with the membranes (5), washed at 60°C to high stringency (0.1% SSC/0.1% SDS), and exposed to Kodak Xomat film at Ϫ70°C. The tWAP mRNA levels were quantitated by slot blot analysis (22). The membranes were hybridized with the tWAP cDNA and washed to high stringency, and each individual slot was excised and placed into 5 ml of organic scintillant.…”

The mammary gland-specific marsupial ELP and eutherian CTI share a common ancestral gene

“…The tammar ELP Rsa I digested LGB (to detect both LGB transcripts [96]) and CST3 probes were either amplified by RT-RCR from tammar mammary gland total RNA or sourced from clones in a tammar mammary gland EST library held by the Cooperative Research Centre for Innovative Dairy Products [19], with plasmid DNA isolated and the cDNA insert amplified by PCR. Membranes were washed (0.1X SSC, 0.1% SDS) twice for 15 min at 60°C, wrapped in cling film, sealed into plastic pockets and exposed to a General Purpose Storage Phosphor screen and scanned on a Typhoon 8600 Scanner (Molecular Dynamics/GE Healthcare).…”

Phylogenetic Analysis of Three Lipocalin-Like Proteins Present in the Milk of Trichosurus vulpecula (Phalangeridae, Marsupialia)

“…The screening was performed according to Sambrook et al (1992). ␤-Lactoglobulin and late-lactation protein clones were isolated using [ 32 P]-dCTPlabeled M. eugenii ␤-lactoglobulin and late-lactation protein cDNA probes (Collet et al, 1989(Collet et al, , 1991. Trichosurin clones were isolated by random selection of phagemid plaques that gave a strong signal when hybridized with ␣[ 32 P]-dCTP-labeled total T. vulpecula cDNA probe but did not hybridize with other possum milk protein cDNAs.…”

“…It is also found in the milk of nonruminant eutherians and has been identified in the milk of two macropod marsupials, Macropus giganteus, the eastern grey kangaroo (Godovac-Zimmermann and Shaw 1987), and Macropus eugenii, the tammar wallaby (Collet et al 1991). ␤-Lactoglobulin is thought to transport retinol and/or fatty acids (Godovac-Zimmermann 1988;Flower 1996).…”