Bidirectional imprinting of a single gene: GNAS1 encodes maternally, paternally, and biallelically derived proteins

Abstract

The GNAS1 gene encodes the ␣ subunit of the guanine nucleotide-binding protein G s , which couples signaling through peptide hormone receptors to cAMP generation. GNAS1 mutations underlie the hormone resistance syndrome pseudohypoparathyroidism type Ia (PHP-Ia), so the maternal inheritance displayed by PHP-Ia has raised suspicions that GNAS1 is imprinted. Despite this suggestion, in most tissues G s ␣ is biallelically encoded. In contrast, the large G protein XL␣s, also encoded by GNAS1, is paternally derived. Because the inheritance of PHP-Ia predicts the existence of maternally, rather than paternally, expressed transcripts, we have investigated the allelic origin of other mRNAs derived from GNAS1. We find this gene to be remarkable in the complexity of its allele-specific regulation. Two upstream promoters, each associated with a large coding exon, lie only 11 kb apart, yet show opposite patterns of allele-specific methylation and monoallelic transcription. The more 5 of these exons encodes the neuroendocrine secretory protein NESP55, which is expressed exclusively from the maternal allele. The NESP55 exon is 11 kb 5 to the paternally expressed XL␣s exon. The transcripts from these two promoters both splice onto GNAS1 exon 2, yet share no coding sequences. Despite their structural unrelatedness, the encoded proteins, of opposite allelic origin, both have been implicated in regulated secretion in neuroendocrine tissues. Remarkably, maternally (NESP55), paternally (XL␣s), and biallelically (G s ␣) derived proteins all are produced by different patterns of promoter use and alternative splicing of GNAS1, a gene showing simultaneous imprinting in both the paternal and maternal directions.

Alternative transcripts and evidence of imprinting of GNAL on 18p11.2

“…As the gene structure of GNAL displays extensive similarity to that of GNAS, a complex imprinted locus, 14 we sought to determine whether the GNAL locus is also imprinted. Genomic imprinting involves allele-specific methylation of CpG island regions, defined here as a sequence of at least 400 bp in length with a G þ C content of at least 50% and an observed:expected ratio of CG dinucleotides greater than 0.6.…”

Diverse splicing mechanisms fuse the evolutionarily conserved bicistronic MOCS1A and MOCS1B open reading frames

“…(Legend on facing page.) may help to select the appropriate initiating methionine but does not encode, and therefore cannot supply, an initiating methionine (Blumenthal, 1995); therefore, the initiating methionine must be intrinsically provided by the MOCS1B ORF+ Because the invertebrate MOCS1B ORFs lack a candidate initiating methionine that would produce a protein including the presumably essential residues, it is highly unlikely that functional MOCS1B protein can be independently translated in these species+ Though modestly transcribed, Northern analyses in human (Reiss et al+, 1998b), as well as in mouse, opossum, and chicken (Fig+ 3) show a single transcript size consistent with mRNAs containing both ORFs, but not smaller transcripts possibly encoding either MOCS1 subunit alone+ It is therefore likely that in vertebrates, as in invertebrates, that MOCS1B is not independently translated because (1) there is no evidence for a small, MOCS1B-specific transcript in four diverse vertebrates, (2) there is no conserved MOCS1B translation initiation codon (see above), and (3) the conventional scanning ribosome translational model (reviewed in Kozak, 1999) only predicts efficient translation of MOCS1A but not MOCS1B from the bicistronic transcripts+ Consequently, we therefore hypothesize that the only protein translated from the bicistronic Type I splice form is monofunctional MOCS1A protein+ Evidence that MOCS1A protein is indeed translated from the bicistronic (Type I) transcript can be seen by noting that the codons specific to this splice form are particularly well conserved (e+g+, the Gly-Gly C-terminal dipeptide; Fig+ 2C and see below), attesting to their protein-coding function+ We envision that the MOCS1B ORF of the bicistronic transcript constitutes an extended 39 UTR, similar to NESP55-GNAS1 transcripts that are thought to produce only NESP55 protein but not G s a due to the lack of an initiating methionine for the latter ORF (Hayward et al+, 1998;Ischia et al+, 1997;Peters et al+, 1999)+ In conclusion, because phylogenetic (this paper) and human mutation (Reiss et al+, 1998b) data imply that MOCS1B protein must be produced from this locus, we propose that the MOCS1B ORF is only translated from the no-nonsense transcripts (Types II-V) as part of a fused MOCS1A-MOCS1B multifunctional protein (Fig+ 2B)+ Recent studies indicate that gene fusion to form multidomain proteins is relatively common in evolution (Doolittle, 1999;Enright et al+, 1999;Marcotte et al+, 1999aMarcotte et al+, , 1999b, including in other eukaryotic genes involved in Mo metabolism )+ Gene fusion begins with a recombination event juxtaposing two ORFs such that transcription of the upstream ORF continues uninterrupted through the downstream ORF+ Should Figure 1+ The Mmus panel shows the patterns from a variety of tissues: B: brain, H: heart, L: liver, and T: testis+ B: Schematic representation of mRNA splice forms and their respective putative protein products+ Genomic organization of human (exons 8-10), fruit fly (exons 3 and 4), and nematode (exons 5-7) are shown at left+ Green shading indicates MOCS1A sequences and blue designates MOCS1B+ The red vertical line represents a nonsense codon that terminates the MOCS1A ORF+ Roman numerals (I-III) in human designate splice donor choices into exon 10; numer...…”

Change in maternal environment induced by cross-fostering alters genetic and epigenetic effects on complex traits in mice

“…Briefly, genomic imprinting refers to parent-of-origindependent gene expression, where the paternally and maternally inherited alleles are expressed to different degrees (Hayward et al 1998;Reik & Walter 2001). This epigenetic effect is primarily caused by differential methylation of the two alleles although histone modifications and microRNA may also modify allele-specific expression (Wilkinson et al 2007;Seitz et al 2008).…”