Related Experiment Video
Updated: Aug 8, 2026
IP-FCM: Immunoprecipitation Detected by Flow Cytometry
Published on: December 2, 2010
Characterization of novel FcepsilonRII/CD23 isoforms lacking the transmembrane (TM) segment in human cell lines
1Institute for Virus Research, Kyoto University, 53 Kawahara-cho, Sakyo-ku, Kyoto, Japan.
Insights
New CD23 transcripts (a' and b') are translated into 40 kDa proteins in human lymphoid cells. These cytoplasmic CD23 proteins may regulate full-length CD23 expression on the cell surface.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- Human Fc epsilon RII/CD23 is a type II transmembrane glycoprotein with two isoforms (a and b).
- Previously identified CD23 transcripts (a' and b') lack exon 3, affecting transmembrane and cytoplasmic domains.
- The protein products of these truncated transcripts were not well characterized.
Purpose of the Study:
- To analyze the protein products of CD23a' and CD23b' transcripts.
- To characterize the expression and potential function of these novel CD23 protein forms.
Main Methods:
- Utilized rabbit polyclonal antibodies against novel amino acid sequences of CD23a' and CD23b'.
- Performed Western blots on COS cells transfected with CD23a' or CD23b' cDNA.
- Conducted in vitro translation assays and analyzed protein glycosylation resistance (Endo H(f), PNGase F).
Main Results:
- CD23a' and CD23b' transcripts were translated into approximately 40 kDa protein molecules.
- These 40 kDa proteins were recognized by antibodies specific to CD23a'/b' and a soluble CD23 fragment antibody.
- Expressed protein products in human cells were specifically recognized by anti-CD23a' and anti-CD23b' antibodies.
- CD23a' and CD23b' molecules were resistant to deglycosylation, indicating post-translational modifications.
Conclusions:
- The a' and b' CD23 transcripts are expressed and translated in human lymphoid cells.
- The resulting 40 kDa cytoplasmic CD23 proteins may have a unique regulatory role in cell surface CD23 expression.
Abstract:
Human FcepsilonRII/CD23 is an approximately 45 kDa type II transmembrane glycoprotein belonging to the C-type animal-lectin family, and has two isoforms (a and b) that only differ in their intracytoplasmic tails. We previously found that in several human and mouse cell lines there were two additional CD23 transcripts (a' and b') lacking the exon 3 that encodes the entire transmembrane segment and a part of cytoplasmic tails. In this study, we analyzed the putative CD23a' and CD23b' products at protein levels and characterized with rabbit polyclonal antibodies against novel amino-acid sequences of the putative CD23a' and CD23b' molecules (anti-CD23a' Ab, anti-CD23b' Ab). Western blots in COS cells transfected with CD23a' or CD23b' cDNA as well as in vitro translation assays showed that the a' and b' CD23 transcripts were translated to about 40 kDa molecules. These 40 kDa molecules were also recognized by a polyclonal antibody against 25 kDa soluble fragment of human CD23. We also found that human cells having mRNAs for CD23a' and CD23b' expressed protein products recognized specifically by anti-CD23a' or anti-CD23b' Ab, respectively. In addition, the CD23a' and CD23b' molecules in transfected COS cells were resistant to Endo H(f) and PNGase F, although these truncated forms as well as the membrane-associated forms had an asparagine residue responsible for the N-linked glycosylation. Taken together, our results show that the a' and b' CD23 transcripts are expressed and translated in human lymphoid cells and that their translated products are retained in the cytoplasm where they might play an unique regulatory role in the expression of the full-length CD23 on the cell surface.
