Luteinizing Hormone and Follicle-Stimulating Hormone Exhibit Different Secretion Patterns from Cultured Madin-Darby Canine Kidney Cells

Jablonka‐Shariff, Albina; Boime, Irving

doi:10.1095/biolreprod.103.022699

Cited by 16 publications

(11 citation statements)

References 51 publications

(87 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The secretion of the heterodimeric gonadotropins from transfected MDCK cells mirrors that seen for the corresponding ␤-subunit (24,25), and the different in vitro profiles recapitulate the exit compartment of LH and CG from the pituitary and the placenta, respectively (19,22,24). The secretion of the hCG␤boCTP chimera from the polarized MDCK cells was predominantly basolateral.…”

Section: Discussionmentioning

confidence: 66%

“…To ensure the fidelity of the LH␤ and CG␤ constructs, both DNA strands were sequenced with an ABI prism DNA sequencer (PerkinElmer Life Sciences). A silent mutation in the Val 25 codon was detected in the cDNA encoding the LH␤CTP variant and was not corrected.…”

Section: Construction Of the Hcg␤-boctp Chimeramentioning

confidence: 99%

“…35 S]cysteine (25 Ci/ ml) for 16 h as described (25). Samples of apical and basolateral media were immunoprecipitated with hCG␤ antiserum, and the reduced proteins were resolved on 15% SDS-PAGE and autoradiographed.…”

Section: Secretion From Mdck Cellsmentioning

confidence: 99%

“…Because of this function, a key question is whether the hCG␤boCTP chimera is sorted to the apical secretory pathway, like the hCG␤ subunit, or is released through the basolateral compartment as the human LH␤ subunit and the O-deglycosylated CG␤ subunit (24). We addressed this point, using the MDCK cell model that was validated previously, to study the secretion patterns of the gonadotropins and their subunits (24,25). The hCG␤boCTP chimera was stably transfected into the cell line, and several clones were selected, grown in Transwells, labeled, and analyzed.…”

Section: N-linked Glycan Modification Of the Lh␤ Subunit Derivates-n-mentioning

confidence: 99%

“…Recent studies, utilizing the Madin-Darby canine kidney (MDCK) model, identified sorting determinants in the ␤-subunit of the hormones (24,25). Whereas the secretion of the hLH␤ subunit from transfected MDCK cells was basolateral, the hCG␤ subunit was routed to the apical secretion pathway, primarily because of the O-glycosylated CTP.…”

mentioning

confidence: 99%

See 4 more Smart Citations

The LHβ Gene of Several Mammals Embeds a Carboxyl-terminal Peptide-like Sequence Revealing a Critical Role for Mucin Oligosaccharides in the Evolution of Lutropin to Chorionic Gonadotropin in the Animal Phyla

Nakav

Jablonka‐Shariff

Kaner

et al. 2005

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

The expression of a previously untranslated carboxylterminal sequence is associated with the ancestral lutropin (LH) ␤ to the ␤-subunit gene evolution of choriogonadotropins (CG). The peptide extension (denoted as CTP) is rich in mucin-type O-glycans and confers new hormonal properties on CG relative to the LH. Although the LH␤ gene is conserved among mammals and only a few frameshift mutations account for the extension, it is merely seen in primates and equids. Bioinformatics identified a CTP-like sequence that is encrypted in the LH␤ gene of several mammalian species but not in birds, amphibians, or fish. We then examined whether or not decoding of the cryptic CTP in the bovine LH␤ gene (boCTP) would be sufficient to generate the LH␤ species of a ruminant with properties typical to the CG␤ subunit. The mutated bovine LH␤-boCTP subunit was expressed and N-glycosylated in transfected Chinese hamster ovary cells. However, unlike human (h) CG␤ CTP, the cryptic boCTP was devoid of mucin O-glycans. This deficiency was further confirmed when the boCTP domain was substituted for the natural CTP in the human CG␤ subunit. Moreover, when expressed in polarized Madin-Darby canine kidney cells, this hCG␤-boCTP chimera was secreted basolaterally rather than from the apical compartment, which is the route of the wild type hCG␤ subunit, a sorting function attributed to the Oglycans attached to the CTP. This result shows that the cryptic peptide does not orientate CG to the apical face of the placenta, to the maternal circulation as seen in primates. The absence of this function, which distinguishes CG from LH, provides an explanation as to why the LH␤ to CG␤ evolution did not occur in ruminants. We propose that in primates and equids, further natural mutations in the progenitor LH␤ gene resulted in the efficient O-glycosylation of the CTP, thus favoring the retention of an elongated reading frame.The family of glycoprotein hormones includes lutropin (LH), 1 follitropin, and thyrotropin, as expressed by the pituitary, and chorionic gonadotropin (CG), which is produced by the placenta of primates and equids (1). Each hormone is a noncovalent heterodimer composed of a common ␣-subunit and a unique ␤-subunit that confers receptor specificity. The ␤-subunits have substantial sequence similarities, including conserved location of cysteine residues that suggests an origin from a common ancestral gene (2). The human (h) CG␤ genes have evolved from an ancestral LH␤ gene by frameshift mutations that resulted in a readthrough into a previously untranslated region and the extension of the reading frame (3, 4) (Fig. 1A). As a result, a short carboxyl-terminal sequence of hLH␤ is replaced by a longer carboxyl-terminal peptide (CTP) in hCG␤ (3-6). In the equine (e), both the placental CG␤ and pituitary LH␤ subunits are products of the same gene (eLH/CG␤) and contain a CTP that is presumed to have evolved by a distinct mechanism (7) (for further explanations see "Results"). Except for primates and equids, no other identified animal species b...

show abstract