Lineage-specific alternative splicing of the human Fc gamma RIIA transmembrane exon requires sequences near the 3' splice site.
The human Fc gamma RIIA gene produces multiple transcripts, including those with (Fc gamma RIIa1) and without (Fc gamma RIIa2) the single exon encoding the transmembrane domain (TM). Previously, a fluorescence-based RT-PCR assay showed lineage-specific differences in Fc gamma RIIA transcript ratios (Fc gamma RIIa2/Fc gamma RIIa1). The mechanism of this lineage-specific expression was investigated in this study. Differential transcript stability does not play a major role, because transcript ratios remained constant in cells with both low (K562) and high (Dami) ratios following actinomycin D treatment. Transient expression studies in K562 and Dami cells using a minigene construct containing a 5.0 kb genomic fragment including the TM exon and adjacent intron and exon sequences showed recapitulation of endogenous transcript ratios. The TM exon was efficiently spliced in by the constitutive splicing machinery in HeLa cells, an Fc gamma RIIA-negative cell line. Lineage-specific TM exon skipping was markedly diminished by two independent minigene mutations: a point mutation of the first nucleotide of the TM exon, and a five basepair intronic deletion near a putative branchpoint. These data demonstrate that cis-acting sequences in or near the TM exon 3' splice acceptor site contribute to lineage-specific differences in Fc gamma RIIA transcript ratios.
Keller
M A
MA
Molecular Biology Graduate Group, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.
McKenzie
S E
SE
Cassel
D L
DL
Rappaport
E F
EF
Schwartz
E
E
Surrey
S
S
D017398:Alternative Splicing; D015703:Antigens, CD; D001483:Base Sequence; D015152:Blotting, Northern; D002478:Cells, Cultured; D002874:Chromosome Mapping; D005091:Exons; D005816:Genetic Complementation Test; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis; D017452:Receptors, IgG / Q000235:genetics*; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection
D015703:Antigens, CD; C080568:Fc gamma receptor IIA; D012333:RNA, Messenger; D017452:Receptors, IgG