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Keywords:

  • AC133 antigen;
  • CD133;
  • prominin-1;
  • splice variant;
  • stem cell marker

Abstract

  1. Top of page
  2. Abstract
  3. Acknowledgments
  4. References

Prominin-1 (CD133), a pentaspan membrane glycoprotein that constitutes an important cell surface marker of various, either normal or cancerous, stem cell populations is widely used to isolate or characterize such cells in different systems. Occurring throughout the metazoan evolution with a remarkably conserved genomic organization, it may be expressed as different splice variants with distinctive characteristics. A rational nomenclature has been proposed earlier for their consistent designation across species. Although generally accepted, it seems to be misunderstood in view of the recent report of novel prominin-1 complementary DNAs in rhesus monkey and humans with improper naming. As this may lead to confusion, we have reexamined the genomic organization of prominin-1 in various primates to provide an update that should further clarify the rationale of the nomenclature for prominin-1 gene products. This report comprises (i) the determination of the genomic organization of prominin-1 gene in two non-human primates, i.e. Macaca mulatta and Pan troglodytes, commonly used in research, (ii) the mapping of a new exon that creates an alternative cytoplasmic C-terminal end of prominin-1, (iii) the identification of various potential PDZ-binding domains generated by alternative cytoplasmic C-terminal tails, suggesting that different prominin-1 splice variants might interact with distinct protein partners, and (iv) a summing up of the different prominin-1 splice variants.

In the article entitled ‘Isolation, molecular cloning and in vitro expression of rhesus monkey (Macaca mulatta) prominin-1.s1 complementary DNA (cDNA) encoding a potential hematopoietic stem cell antigen’, recently published in Tissue Antigens by Husain et al. (1), the authors refer to a previous article from our group proposing a unifying nomenclature for the designation of the prominin family gene products (2) based on their conserved genomic organization across species (3, 4). The aim of this nomenclature was to attribute the same designation to a given splice variant irrespective of the species as splice variants described in one organism could be predicted to exist in others, with similar characteristics at least among mammals. This nomenclature seems to be generally accepted in the field (5–9), but the rationale underlying it may remain unclear as the rhesus prominin-1 splice variant described by Husain et al. does not correspond to the s1 splice variant (1), and several unpublished human prominin-1 splice variants appear in the NCBI GenBank database (accession numbers AY449690 to AY449693) with the incorrect suffix according to this same nomenclature. Given the wide use of prominin-1 (also termed CD133) for the characterization of stem and progenitor cell populations in different normal tissues (10–13) as well as in cancers [(14–16); for review, see (17)], it is important to clarify the rationale of this nomenclature. We believe that it is essential to maintain a consistent designation with regard to potential cross-specificity toward particular epitopes, e.g. stem-cell-associated AC133 epitope (18). Moreover, when dissecting the differential tissue expression and function of these prominin-1 splice variants, it is important to refer to homologous molecules in the different model organisms. Anarchic reference to the nomenclature can only bring in confusion as to the interpretation of data generated in different models. The chronology of the identification of prominin-1 splice variants in one species or the other is not necessarily related to the predominance of one variant in one species compared with the others and therefore cannot serve as a base for their designation. For instance, prominin-1.s1 was first described in mouse then in humans, while prominin-1.s2 was first described in human then in mouse, or in the same line, only prominin-1.s1 and s7 have been described in rat (19). Cross-species sequence comparison should be made between homologous variants in order to be significant. We therefore propose to sum up and update the nomenclature of the different prominin-1 splice variants and the underlying genomic features in humans and different animal models.

The prominin-1 gene is located on chromosome 4 in humans and chromosome 5 in mice and spans more than 150 kb (3, 20, 21). Its genomic structure, i.e. exon/intron boundaries, is strikingly similar across species (3). Since the original description of prominin-1 (22), several splice variants affecting the open reading frame have been identified in mouse (2, 23, 24) and in humans (25, 26) and their expression was characterized (4). Factors regulating the in vivo expression of prominin-1 remain to be determined, but the messenger RNA profile suggests that the mouse prominin-1 splice variants are tissue specific and developmentally regulated (4).

To date, alternative splicing was found to affect the N-terminal domain, the first and second extracellular loops or mostly the cytoplasmic C-terminal domain, which might implicate distinct cytoplasmic protein-interacting partners. The splice variants s1 and s3–s8 were first isolated in mouse and defined four alternative C-termini for prominin-1, which occur through (i) intron retention, (ii) exon skipping, or (iii) usage of a cryptic acceptor site (4). This splicing cassette, which locates within a cluster of short exons and introns, is conserved across species allowing the prediction of similar splice variants (4). Additional splice variants that define two more alternative C-termini have been isolated in humans (AY449689, AY449691, and AY449693). By comparing the prominin-1 cDNA sequence (nucleotides 2463–2487) isolated from KG-1a cell line (see AY449689) with the human prominin-1 genomic sequence (NC_000004), we have identified a novel facultative exon (SSWVTSVQ), i.e. exon 25, within this splicing cassette (positions 15591022 to 15590998 of the genomic clone) that conforms to the consensus GT-AG rule and encodes residue 822–829 of this particular prominin-1 variant (from here on referred to as s9) (Tables 1 and 2). Importantly, exon 25 being 25 nucleotides long, its insertion induces a frameshift on the following exon 26b, and hence, a premature ending following a C-terminal cysteine (residue 830) compared with the s1 and s2 splice variants (Table 3).

Table 1.  Genomic structure of primate prominin-1 genesa
ExonHuman cDNA (AF027208), 3794 ntHuman cDNA (AY449689), 2493 ntHuman chromosome 4 (NC_000004)Rhesus cDNA (AY903606), 3927 ntRhesus chromosome 5 (NC_007862)Chimpanzee predicted (XM_517115)Chimpanzee chromosome 4 (NC_006471)
  • cDNA, complementary DNA; ND, not determined; Nt, nucleotide; —, absence of the exon in the cDNA under consideration.

  • a

     The numbering of the exons begins with the exon bearing the initial start codon. The 5′ regions (UTRs) were ignored for the sake of simplicity in comparing the different gene sequences. Exon 29 contains the 3′ UTR with the polyadenylation sequence. The exon–intron structure of primate prominin-1 genes were completed by comparing the sequences of the humans, rhesus, and chimpanzee genomic DNA (accession numbers: NC_000004, NC_007862 and NC_006471, respectively) with that of human prominin-1.s2 and −s9 cDNAs (AF027208 and AY449689, respectively) and that of mouse prominin-1.s3 and −s8 cDNAs (AF305215 and BC028286, respectively). Nucleotide and protein databases were searched at the National Center for Biotechnology Information (NCBI) using the BLAST network services.

  • b

     Exon 19 has been only identified in murine prominin-1.s8 so far (see Table 2).

  • c

     Exons 26a and 26b are created by alternative splice acceptor sites, and they are initially referred as exons 25a and b in a previous publication (4).

11–2571–22015686664–156864085–44010892907–10892472308–73916021345–16020914
2258–313221–27615649722–15649667441–49610853557–10853502740–79515984217–15984162
3314–34015646482–15646456497–52310850408–10850382796–82215980970–15980944
4341–546277–48215644230–15644025524–72910848142–10847937823–102815978717–15978512
5547–667483–60315636033–15635913730–85010839541–108394211029–114915970521–15970401
6668–731604–66715635079–15635016851–91410838634–108385711150–121315969576–15969513
7732–821668–75715634136–15634047915–100410837679–108375901214–130315968613–15968524
8822–1039758–97515629261–156290441005–122210832868–108326511304–152115962399–15962182
91040–1114976–105015626961–156268871223–129710830557–108304831522–159615960098–15960024
101115–11781051–111415624059–156239961298–136110827613–108275501597–166015957207–15957144
111179–13381115–127415619829–156196701362–152110823685–108235261661–182015952979–15952820
121339–14911275–142715617411–156172591522–167410820925–108207731821–197315950562–15950410
131492–16151428–155115611340–156112171675–179810814875–108147521974–209715944639–15944516
141616–17191552–165515609209–156091061799–190210813072–108129692098–220115942516–15942413
151720–18041656–174015604792–156047081903–198710808780–108086962202–228615938123–15938039
161805–19481741–188415603112–156029691988–213110807105–108069622287–243015936442–15936299
171949–20201885–195615602014–156019432132–220310806037–108059662431–250215935353–15935282
182021–21131957–204915600545–156004532204–229610804562–108044702503–259515933920–15933828
19b ND ND ND
202114–21672050–210315598437–155983842297–235010802763–108027102596–264915931721–15931668
212168–22482104–218415596758–155966782351–243110801104–108010242650–273015930033–15929953
222249–23172185–225315596549–155964812432–250010800895–108008272731–279915929824–15929756
232318–24102254–234615595076–155949842501–259310799403–107993112800–289215928351–15928259
242411–25262347–246215591258–155911432594–270910795923–107958082893–300815924544–15924429
252463–248715591022–1559099810795690–1079566615924302–15924278
26ac15590886–1559084610795553–1079551315924165–15924125
26bc2527–25502488–249315590625–155906022710–273310795292–107952693009–303215923905–15923882
272551–2619 15590184–1559011610794849–107947813033–310115923464–15923396
282620–2659 15581803–155817642734–277010786491–107864553102–314115917934–15917895
292660–3794 15580089–155789552771–390410784777–107836473142–428415916215–15915073
Table 2.  Presence or absence of facultative exons in various prominin-1 splice variantsa
Splice variantStructure (facultative exons included in the coding sequences)Mus musculusRattus norvegicusHomo sapiensMacaca mulattaPan troglodytes
39192526a26b2728
  • a

     The structure of the different prominin-1 splice variants is indicated by the presence (+) or absence (−) of the facultative exons numbered according to Table 1. Prominin-1 splice variants identified in each species are indicated by their respective GenBank accession number and protein sequence length (number of amino acid residue).

s1++++AF026269 (858)AF386758 (857)AF507034 (856) 
s2+++++AF039663 (867) AF027208 (865)XM_001100223 (864)XM_517115 (865)
s3++AF305215 (834) 
s4+AY223521 (804) 
s5+AY223522 (809) 
s6+AY099088 (823) 
s7++AK029921 (827)AY262731 (826)AY449690 (825) 
s8++++BC028286 (842) 
s9+++ AY449689 (830) 
s10+++ AY449691 (833) 
s11+++ AY449692 (834) 
s12++++ AY449693 (842)AY903606 (841) 
Table 3.  Alternative prominin-1 C-terminia
  • cDNA, complementary DNA; C, chimpanzee; H, human; R, rhesus.

  • a 

    Alternative splicing might generate six distinct cytoplasmic C-terminal tails of prominin-1. Sequences from isolated cDNAs (see Table 2) appear in regular characters, while predicted sequences are given in italics. Exons 25, 26a, 26b, and 27 are given in cyan, red, green, and blue, respectively. Asterisks indicate exon 25 being 25 nucleotides long introduces a frameshift on the following exon 26b. Nonconserved amino acid residues within the rhesus sequence are given in bold underlined.

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Thus, the prominin-1 coding region spans 28 exons among which 7 are facultative with exon 26 bearing two mutually exclusive acceptor sites a and b (Table 2). Remarkably, we have identified a nearly identical genomic structure for two others primates, i.e. rhesus and chimpanzee (on chromosomes 5 and 4, respectively) by similar sequence comparison (Tables 1 and 3), lending further weight to the rationale underlying the nomenclature for prominin-1 gene products. Hence, prominin-1 from chimpanzee is predicted to display the characteristic five transmembrane domains (see updated sequence XM_517115) like in the other species, and the s1 variant would be predicted to contain 865 residues, contrarily to what appears in table 1 of Husain et al. (1). Together, 12 different prominin-1 splice variants affecting the protein sequence have been described to date in rodents and primates (Table 2). Some of them, e.g. s1, s2, s7, s12, have been detected in several species. Completing the chart for a given species might thus only be a question of time.

It appears therefore that a prominin-1 splice variant with a cytoplasmic C-terminal tail that is 24 amino acid shorter than the human prominin-1.s2 (AF027208) like the rhesus prominin-1.s1 splice variant described by Husain and colleagues does exist in humans. Consequently, this rhesus prominin-1 splice variant aligns better with the 842 amino acid long human splice variant encoded by an unpublished cDNA sequence deposited in GenBank (accession number AY449693; here referred to as s12; Table 2) than with human prominin-1.s2, yielding 95.7% amino acid sequence identity rather than 93.2% (using the stretcher program from the EMBOSS package with a EBLOSUM 62 matrix and gap and extend penalty of 12 and 2, respectively). The prominin-1 sequence from Macaca mulatta (AY903606) has therefore been renamed prominin-1.s12 (S. M. Husain, personal communication, St. Jude Children’s Research Hospital, Memphis, TN).

The importance of using the same suffix to design a given splice variant irrespective of the species is best illustrated when considering the cytoplasmic C-terminal tail of prominin-1 (Table 3). Although no information is currently available as to potential cytoplasmic proteins interacting with prominin-1, the presence of distinct C-terminal domains suggests that different prominin-1 splice variants might interact with alternative protein partners. Interestingly, we found that the last four C-terminal amino acids of the s3, s4 and s5 variants, i.e. HFTL, exhibit the characteristics of a class II PDZ-binding domain with a hydrophobic residue in position 0 and −2 (X-φ-X-φ; X, unspecified amino acid and φ, hydrophobic residue), while the C-terminus of the s9 splice variant, i.e. SVQC, would be highly related to a class III PDZ-binding domain (X-X-C) and rodent s1, s2, s7, s8 and s11 (PSQR) to a class I PDZ-binding domain (X-S/T-X-φ) (27, 28). In keeping with a potential interaction with PDZ-domain-containing protein, we have recently found that prominin-2, the prominin-1 paralogue (3), binds to a novel splice variant of the glutamate receptor-interacting protein, a PDZ-domain-containing protein in a yeast two-hybrid screening (Kathrin Opherk and DC, unpublished data; see GenBank accession number AY255674). Further studies are needed to determine whether prominin-1 molecules interact with various PDZ-domain-containing proteins as well as to study the physiological relevance of such interaction.

In conclusion, we have shown (i) that the genomic organization of prominin-1 gene in two non-human primates is highly similar to that in the human counterpart and (ii) the existence of a novel conserved facultative exon that creates an alternative cytoplasmic C-terminal end of prominin-1. Therefore, we recommend that the designation of the prominin-1 splice variants from Homo sapiens (AY449689 to AY449693) be corrected according to the present Table 2 in order to conform to the nomenclature for prominin-1 gene products they appear to refer to.

Acknowledgments

  1. Top of page
  2. Abstract
  3. Acknowledgments
  4. References

WBH was supported by the Deutsche Forschungsgemeinschaft (SPP 1109, Hu275/7-3; SPP 1111, Hu275/8-3; SFB/TR13-04 B1; SFB 655 A2) and DC by the Deutsche Forschungsgemeinschaft (SPP 1109, CO 298/2-2; SFB/TR13-04 B1; SFB 655 A13) and Sächsisches Ministerium für Wissenschaft und Kunst–Europäischer Fond für Regionale Entwicklung (4212/05-16).

References

  1. Top of page
  2. Abstract
  3. Acknowledgments
  4. References
  • 1
    Husain SM, Shou Y, Sorrentino BP, Handgretinger R. Isolation, molecular cloning and in vitro expression of rhesus monkey (Macaca mulatta) prominin-1.s1 complementary DNA encoding a potential hematopoietic stem cell antigen. Tissue Antigens 2006: 68: 31724.
  • 2
    Fargeas CA, Corbeil D, Huttner WB. AC133 antigen, CD133, prominin-1, prominin-2, etc.: prominin family gene products in need of a rational nomenclature. Stem Cells 2003: 21: 5068.
  • 3
    Fargeas CA, Florek M, Huttner WB, Corbeil D. Characterization of prominin-2, a new member of the prominin family of pentaspan membrane glycoproteins. J Biol Chem 2003: 278: 858696.
  • 4
    Fargeas CA, Joester A, Missol-Kolka E, Hellwig A, Huttner WB, Corbeil D. Identification of novel prominin-1/CD133 splice variants with alternative C-termini and their expression in epididymis and testis. J Cell Sci 2004: 117: 430111.
  • 5
    Shmelkov SV, St Clair R, Lyden D, Rafii S. AC133/CD133/prominin-1. Int J Biochem Cell Biol 2005: 37: 7159.
  • 6
    Zhang CC, Lodish HF. Murine hematopoietic stem cells change their surface phenotype during ex vivo expansion. Blood 2005: 105: 431420.
  • 7
    Hess DA, Meyerrose TE, Wirthlin L et al. Functional characterization of highly purified human hematopoietic repopulating cells isolated according to aldehyde dehydrogenase activity. Blood 2004: 104:164855.
  • 8
    Wu X, Rabkin-Aikawa E, Guleserian KJ et al. Tissue-engineered microvessels on three-dimensional biodegradable scaffolds using human endothelial progenitor cells. Am J Physiol Heart Circ Physiol 2004: 287: H4807.
  • 9
    Yu Y, Flint AF, Mulliken JB, Wu JK, Bischoff J. Endothelial progenitor cells in infantile hemangioma. Blood 2004: 103: 13735.
  • 10
    Yin AH, Miraglia S, Zanjani ED et al. AC133, a novel marker for human hematopoietic stem and progenitor cells. Blood 1997: 90: 500212.
  • 11
    Uchida N, Buck DW, He D et al. Direct isolation of human central nervous system stem cells. Proc Natl Acad Sci U S A 2000: 97: 147205.
  • 12
    Richardson GD, Robson CN, Lang SH, Neal DE, Maitland NJ, Collins AT. CD133, a novel marker for human prostatic epithelial stem cells. J Cell Sci 2004: 117: 353945.
  • 13
    Lee A, Kessler JD, Read T-A et al. Isolation of neural stem cells from postnatal cerebellum. Nat Neurosci 2005: 8: 7239.
  • 14
    Singh SK, Hawkins C, Clarke ID et al. Identification of human brain tumour initiating cells. Nature 2004: 432: 396401.
  • 15
    O’brien CA, Pollett A, Gallinger S, Dick JE. A human colon cancer cell capable of initiating tumour growth in immunodeficient mice. Nature 2007: 445: 10610.
  • 16
    Ricci-Vitiani L, Lombardi DG, Pilozzi E et al. Identification and expansion of human colon-cancer-initiating cells. Nature 2007: 445: 1115.
  • 17
    Fargeas CA, Fonseca AV, Huttner WB, Corbeil D. Prominin-1 (CD133): from progenitor cells to human diseases. Future Lipidol 2006: 1: 21325.
  • 18
    Florek M, Haase M, Marzesco A-M et al. Prominin-1/CD133, a neural and hematopoietic stem cell marker, is expressed in adult human differentiated cells and certain types of kidney cancer. Cell Tissue Res 2005: 319: 1526.
  • 19
    Corbeil D, Fargeas CA, Huttner WB. Rat prominin, like its mouse and human orthologues, is a pentaspan membrane glycoprotein. Biochem Biophys Res Commun 2001: 285: 93944.
  • 20
    Maw MA, Corbeil D, Koch J et al. A frameshift mutation in prominin (mouse)-like 1 causes human retinal degeneration. Hum Mol Genet 2000: 9: 2734.
  • 21
    Shmelkov SV, Jun L, St Clair R et al. Alternative promoters regulate transcription of the gene that encodes stem cell surface protein AC133. Blood 2004: 103: 205561.
  • 22
    Weigmann A, Corbeil D, Hellwig A, Huttner WB. Prominin, a novel microvilli-specific polytopic membrane protein of the apical surface of epithelial cells, is targeted to plasmalemmal protrusions of non-epithelial cells. Proc Natl Acad Sci U S A 1997: 94: 1242530.
  • 23
    Miraglia S, Godfrey W, Buck D. A response to AC133 hematopoietic stem cell antigen: human homologue of mouse kidney prominin or distinct member of a novel protein family? Blood 1998: 91: 43901.
  • 24
    Corbeil D, Röper K, Fargeas CA, Joester A, Huttner WB. Prominin: a story of cholesterol, plasma membrane protrusions and human pathology. Traffic 2001: 2: 8291.
  • 25
    Miraglia S, Godfrey W, Yin AH et al. A novel five-transmembrane hematopoietic stem cell antigen: isolation, characterization, and molecular cloning. Blood 1997: 90: 501321.
  • 26
    Yu Y, Flint A, Dvorin EL, Bischoff J. AC133-2, a novel isoform of human AC133 stem cell antigen. J Biol Chem 2002: 277: 207116.
  • 27
    Harris BZ, Lim WA. Mechanism and role of PDZ domains in signaling complex assembly. J Cell Sci 2001: 114: 321931.
  • 28
    Sheng M, Sala C. PDZ domains and the organization of supramolecular complexes. Annu Rev Neurosci 2001: 24: 129.