Proc. Natl. Acad. Sci. USAVol. 84, pp. 9155-9159, December 1987Immunology

Structure and expression of the human and mouse T4 genesPAUL JAY MADDON*, SUSAN M. MOLINEAUXt, DOUGLAS E. MADDONt, KATHRYN A. ZIMMERMAN*,MAURICE GODFREYt, FREDERICK W. ALT*, LEONARD CHESSt, AND RICHARD AXELt*Department of Biochemistry and Molecular Biophysics, tDepartment of Medicine, and tHoward Hughes Medical Institute, College of Physicians andSurgeons, Columbia University, New York, NY 10032

Contributed by Richard Axel, August 21, 1987

ABSTRACT The T4 molecule may serve as a T-cell recep-tor recognizing molecules on the surface of specific target cellsand also serves as the receptor for the human immunodefi-ciency virus. To define the mechanisms of interaction of T4with the surface of antigen-presenting cells as well as withhuman immunodeficiency virus, we have further analyzed thesequence, structure, and expression of the human and mouseT4 genes. T4 consists of an extracellular segment comprised ofa leader sequence followed by four tandem variable-joining(VJ)-like domains, a transmembrane domain, and a cytoplas-mic segment. The structural domains of the T4 protein deducedfrom amino acid sequence are precisely reflected in theintron-exon organization of the gene. Analysis of the expressionof the T4 gene indicates that T4 RNA is expressed not only inT lymphocytes, but in B cells, macrophages, and granulocytes.T4 is also expressed in a developmentally regulated manner inspecific regions of the brain. It is, therefore, possible that T4plays a more general role in mediating cell recognition eventsthat are not restricted to the cellular immune response.

Analysis of the surface glycoproteins of peripheral T lym-phocytes demonstrates that mature T cells segregate into oneoftwo classes: those that express the surface glycoprotein T4(CD4) and those that express the glycoprotein T8 (CD8) (1).The T4 molecule is primarily expressed on helper T lympho-cytes, whereas T8 is expressed on cytotoxic and suppressorT cells (2, 3). T8+ T lymphocytes interact with a broad set oftarget cells that express class I major histocompatibilitycomplex (MHC) gene products whereas T4+ T cells interactwith a more restricted subset of targets, largely macrophagesand B cells, that express class II MHC molecules (2, 3). Thishas led to the suggestion that the specificity of interaction ofsubpopulations of T lymphocytes with various target cellsresults in part from the association of T4 and T8 with theproducts of different MHC genes. T4 may not only serve asa receptor recognizing molecules on the surface of targetcells, but also serves as the receptor for the human immu-nodeficiency virus (HIV) (4-7).We have isolated (8) the cDNA and the gene encoding T4

and have determined the nucleotide sequence of the full-length cDNA clone. To define the mechanisms of interactionofT4 with the surface of both antigen-presenting cells as wellas with HIV, we have further analyzed the sequence,structure, and expression of the human and mouse§ T4 genes.Human T4, as well the mouse homologue L3T4, exhibit apolyimmunoglobulin-like structure with four tandem vari-able-joining (VJ)-like domains. This polyimmunoglobulin-like structure of T4 is homologous to an increasingly largenumber of recognition molecules. Moreover, we observe thatT4 expression is not restricted to T cells, suggesting that T4plays a more general role in cell-cell interactions.

MATERIALS AND METHODS

All materials and procedures have been described (7-9).

RESULTST4 Exhibits a Polyimmunoglobulin-like Structure. The ami-

no acid sequence of human T4 and the murine and rathomologues have been deduced from the nucleotide se-quences (8, 10-12). The human and mouse T4 sequencesdetermined in our laboratory are shown in Fig. 1. Initialsequence analysis demonstrated that the N-terminal 100amino acids share considerable homology with immunoglob-ulin K light-chain V regions (8). Williams and his colleagues(12) have suggested that the extracellular portion of T4consists not of one, but of four, tandem immunoglobulin-likedomains, generating a polyimmunoglobulin-like structure.This suggestion is supported by a comparison of the se-quences of the human and mouse T4 molecules, by homologybetween T4 and a second polyimmunoglobulin structure, thepolymeric immunoglobulin receptor, as well as by the struc-ture of the chromosomal gene encoding T4.Amino acid residues 1-94 of mature T4 and L3T4 share

-35% homology with immunoglobulin K light-chain V do-mains (Fig. 1). This V-like domain contains two cysteineresidues separated by 69 amino acids that form the conservedintrastrand disulfide bond characteristic of immunoglobulinV domains (13). Moreover, 11 of the 14 invariant residuescharacteristic of light-chain V regions are conserved in L3T4.Aside from homologies at the level of individual amino acids,this N-terminal domain shares structural features with im-munoglobulin V regions. Analysis of probable p-strands andp-turns (14) suggests the presence of seven p-strands withinthis V-like domain that closely match those found in immu-noglobulin V domains. Interestingly, the N-terminal V-likedomain is followed by a stretch of amino acid residues thatshares homology with the J region of immunoglobulins andT-cell antigen receptors (Fig. 1) (8).

Analysis of the remaining 250 extracellular amino acidsreveals three additional VJ-like domains (V2J2, V2J3, andV4J4) that share amino acid and structural homology withimmunoglobulin V and J domains but have diverged consid-erably from prototype V and J regions. The second and fourthV-like domains of T4 and L3T4 each contain two disulfide-linked cysteine residues (Fig. 1) (13). The spacing of thecysteines in these two V-like domains (28 residues in V2 and42 residues in V4) is considerably shorter than in prototypeimmunoglobulin V domains. The V3 domain lacks cysteineresidues, but sequence analysis suggests that this domainmaintains the potential to form the seven 8-strands charac-teristic of V domains (Fig. 1). These truncated and highly

Abbreviations: HIV, human immunodeficiency virus; MHC, majorhistocompatibility complex; J, joining; V, variable.§This sequence is being deposited in the EMBL/GenBank data base(Bolt, Beranek, and Newman Laboratories, Cambridge, MA, andEur. Mol. Biol. Lab., Heidelberg) (accession no. J03564).

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The publication costs of this article were defrayed in part by page chargepayment. This article must therefore be hereby marked "advertisement"in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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T4 QN KV|VLGKKDTVELTCTAISQKKISI Q HWNN IIKILGIN QG SFLTK 49L3T4 fKTT LVjLGKEESAMPCESSQKKITVFTWFSDQRKILGQKGVLI RG 50

caa999 egcgctggt9ct999ha"gaa999ha tcaa daactgccctgcga *tWtccc9agaagatgWcac gtc ttcacct99a*attc tctgaccaamaggta m fu ttg

40 V1-.AVVVVMNVV\MA /vVWVVWVM AAAV&V\5VW\ /\VW\A

T4 P[KLN-[ RA YRRS L[DQ GNFPL IIKNLKIEDSDTYICEVED KEEVQLiZQL 98L3T4 GSPSQFDFDSKKGAMEKGSFPLIINKLK | ICE|L[DNRCKEEV[EW 100

ggttcgccettcgcg fi9tgacgt fi9t gutccaaug99ggcutg9g9agaau9gatc9 fi cctctcatc atcm*taacttaagatggpaugactctcaguct tat atc tgtgp9gct9gagaac*9agguauggagg9tggpgftgtggt9

______ V1

MA/W\* /ANW\W ~WWWV\A/VVVVVVV MAV/WVW\AT4 VFGLTANSDTHL SLLQLTLT LES PPGS S SV Q RS PRGNI QRGKT 147L3T4 MKVWFS PGTSL LQGQSLTLT.DSNSK !NLTECKHKKGKVVSGSKVLS 150

JA V2

AAA/V\ NAA~WVVV\A ANVVVVVW/\A ANWAAAV".A ANAAT4 VSQLELQDSGT[TrCTVILQNLQKNKVE FKIDIVVLAFQKASSIVYKKEGEQVE 197L3T4 MSNLRVQDSDFW TVTLDIQK(NW[GMTLS VLGFQ STAITAY SEGESAE 200

utgtcuacctuugfcgacugttcgac99ettctggaactgcccg9tguccctggucc¢agauuaaguctg flcgca9etgacactctcgtgctggg fit mcagga~cugcgtatc 9 * t9"@ucggctaagaggggggcgga

V2 ,-4J2 - V3

/\AVW\A1 V\W \/\ /W/VAW\/A A/V/NWVW/V\,/ V\/V/\\T4 FS 249L3T4 FSFPLNAE E-- NGWGELMJK[ KDSFFQPWI SFS I NKEVSVQKSTK[ LK 250

t tc Cti c t ac fi gcu gg ga Sac gg tg g*a cig atg tgg aa gc gag maa gt tct tC -tc cog ccc tgg at WCttc tc a" aug sec gasgu gtg tCc gtu cam au tC aemcca gac cc aug ctc

/VWWW\ /VWAWVWA/VV\ AANWvWA V A/AV\T4 QMGKK[P H[TLPQALP[Y]AGSGNLTLALEAKTGKLHQEVNLVVMRAT[QLQK 301L3T4 QLKETILPL|TLKI VSLQFAGSGN :LTL D-K-GTLHQEVNLVVM|KVAQENS 300

cagctg au g c" c c ccu ctc acc ctc au ut cc c gtct9C cb cgCg it gct ggt tt ggc mc ct ac ct act tg gC - gs - ggg ace ctg cat cug gea gtg Mac ctg g gtg 9tg a" gtg gct cag ct age amaV3 J3

T4 N LTCEVWGPTSPKLMLSLKL[NKEAK[SKREKAVWVLN[EAGMWQCLLSID 351L3T4 T LTCEVMJGPTSPK MRLTL QENRVSEEQKVQVVAPETGLWQCLLSE 350

act fg accmtgtggtg ggo Ct ace tct Ccceag atgg9 &ccdtac tgaugcgguag Maccogg gca gggtc tcttgog9t99ggaugtu gtt cougtggtgg9Cctgugucug tgtgcg tgt Ctu tg 9tgau

V4

T4 SGQ[LLE[NIK [VPTWSTPVQPM [LIVLGGVAGLUL FIGLGIFFCVRCRHR 402L3T4 GDKVKMDSRIQVLSRGVNQTVFLA-CVLGGSFGFLGFLGLCLC[CVRCRHQ 400

ggt get uag gtc uag utg 9e tcc aOgg tc cug gn tta tcc uag 9g gtg auc cug Ca gtg ntc ctg gct - tgc gtg ctg ggt ggc t Ctcgec ft ctg ggt tc ctt ggg ctc tec utc ctc tgc tgt gtc agg tgc cng cac cm

J4 TMI

V VT4 RRQAERMSQIKRLLSEKKTCQCPHRFQKT TCSPI 435L3T4 QRQAARMSQIKRLLSEKKTCQCPHRMQKSHNLI 433

cog cgc cug gcu Cu cgu ut9 tet coguteaugg ag ctc ctc agt gag uag uug acc tgc cag tgc ccc cac cgg atg cug aug ugc cat eat ctc utc

CYT b

FIG. 1. Alignment of T4 and L3T4 amino acid sequences, L3T4 nucleotide sequence, and position of introns. The numbers on the right areamino acid residue positions. The L3T4 nucleotide sequence appears below the amino acid sequence. The V-like (V), J-like (J), transmembrane(TM), and cytoplasmic (CYT) regions are indicated by horizontal arrows below the L3T4 nucleotide sequence, although the exact boundariesare ambiguous. The six extracellular cysteines are marked above the sequence by circles-the first cysteine in the disulfide linkage by 0 andthe second by o. The residues that are predicted to form 3-strands are marked by sawtooth lines above the sequence (14). The position of intronsis marked by triangles above the sequence. The potential N-linked glycosylation sites (Asn-Xaa-Thr) in L3T4 (four) and T4 (two) appear asstippled boxes in the sequence. The amino acid homologies between individual domains of L3T4 and T4 (8) are as follows: V1J1, 53%; V2J2,50%o; VJ3, 57%; V4J4, 52%; TM, 52%;o and CYT, 80%. The L3T4 cDNA clone pL3T4B was isolated by screening a mouse thymocyte cDNAlibrary with the human T4 cDNA probe pT4B (8).

divergent domains, however, reveal sequence and structural tion of invariant residues about the cysteines and character-homologies with prototype V regions that include conserva- istic hydrophobic stretches that stabilize the antibody fold.

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Interestingly, a functional immunoglobulin V region lackingthe classical disulfide linkage, as in V3, has been described(15). Thus, three of the four extracellular domains in T4 andL3T4 reflect highly divergent members of the immunoglob-ulin gene family. Moreover, as in the case of V1, theremaining V-like domains are each followed by a J-like regionwith various degrees of homology to prototype immunoglob-ulin J regions.The presence of multiple V-like domains in a single

molecule has also been described for the neural cell-adhesionmolecule (16), the myelin-associated glycoprotein (17), andthe polymeric immunoglobulin receptor (18). Interestingly,one of these proteins, the polymeric immunoglobulin recep-tor, is not only structurally homologous to T4 but also sharessignificant sequence homology with T4 (12).The V4J4 domain of T4 and L3T4 is followed by a

hydrophobic transmembrane sequence that is =44% homol-ogous to the transmembrane region of class II MHC P chains(8). This domain of L3T4 is followed by a 38-amino acidcytoplasmic domain consisting of charged residues that differat only eight positions with human T4 (Fig. 1).

Structure of the T4 Gene. The polyimmunoglobulin-likestructure of T4 and L3T4 suggested from the sequence ofcDNA clones is supported by an analysis of the arrangementof introns and exons in the T4 gene. A set of overlappingclones spanning the T4 gene were isolated (8) from humangenomic libraries constructed in the X cloning vectors Charon4A and EMBL-3. Four overlapping clones that encompassthe entire T4 gene were characterized by restriction analysis,Southern blotting, and nucleotide sequencing (8) (Fig. 2). TheT4 gene consists ofnine exons split by eight introns and spansa region of -33 kilobases (kb) (Fig. 2). Nucleotide sequenceof the 5'-donor and 3'-acceptor splice sites in each intronconforms to the GT ... AG rule and adheres closely to theconsensus sequences compiled for exon-intron boundaries(19). Interestingly, each of the structural domains deducedfrom the cDNA sequence (8) are separated from one anotherby an intron (Figs. 1 and 2). Exon 1 encodes the 5'-untranslated region and most of the hydrophobic signalsequence. Exon 2 encodes the last seven amino acids of thesignal sequence and the first 48 amino acids ofthe V1 domain.Exon 3 encodes the second half of the V1 domain as well asthe J1 segment. Thus, the V1 domain is encoded by two exonsseparated by 'w12 kb, suggesting that the primitive immuno-globulin exon may have originated as a half V domain thatduplicated and fused later in evolutionary time.

The three remaining VJ-like regions of the extracellulardomain are each encoded by individual exons (Figs. 1 and 2).The exons encoding the extracellular VJ-like domains arefollowed by an exon that encodes the transmembrane regionand the first six amino acids of the cytoplasmic segment. Thebulk of the cytoplasmic domain is encoded by an exon thatends 12 amino acids from the C terminus. The last and thelargest exon encodes the remaining six amino acids of thecytoplasmic domain and 3'-untranslated DNA. Thus, thestructural domains of the T4 protein are precisely reflected inthe intron-exon organization of the gene.

Expression of T4 mRNA. The polyimmunoglobulin-likestructure of T4 resembles the structure of other recognitionmolecules. T4 is likely to serve as a recognition moleculemediating T-cell-target-cell interaction. The availability ofT4 and L3T4 cDNAs now permits us to examine whether T4expression is restricted to the immune system or whetheradditional cell types also express the T4 molecule. We have,therefore, analyzed the RNA populations of human andmurine tissues, hematopoietic cells, and cell lines for theexpression ofT4 and L3T4RNA byRNA gel blot analysis (8).Examination of several human and mouse tissues reveals thatT4 and L3T4 are expressed in cells ofthe thymus, spleen, andbrain (Figs. 3-5). We have demonstrated (7) that two RNAspecies are observed in human brain: one 3 kb in length thatcorresponds to the mRNA expressed by T lymphocytes, aswell as a shorter, variant RNA 1.8 kb long (Fig. 5). Immu-nohistochemical analyses suggest that surface T4 in the brainis largely restricted to invading macrophages (20). Thus the3-kb mRNA in human brain is likely to derive at least in partfrom macrophages. The variant 1.8-kb RNA, however, hasnever been observed in cells of hematopoietic origin and,therefore, may derive from neurons or supporting cells in thecentral nervous system (Fig. 5) (7, 8). The observation that T4is expressed in the brain is of interest since T4 is the receptorfor HIV, and HIV frequently infects cells within the centralnervous system (4-7, 21).

In contrast, the mouse brain expresses only the shorter,variant form ofL3T4RNA -2.2 kb long (Figs. 3 and 4) (7, 10).RNA gel blot analysis of poly(A)+ RNA from various regionsof the adult mouse forebrain reveals the presence of a 2.2-kbL3T4 RNA in the cortex and striatum, with higher levels inthe striatum (Fig. 4A). No L3T4 RNA is detectable in themidbrain, hippocampus, cerebellum, medulla, and pituitary.We estimate that in the striatum, the level of expression ofL3T4 RNA is lower than in the thymus by a factor of 25. Toexamine the developmental expression of L3T4 in the brain,

I- 1.8 Kb- .-- 20 Kb- D4 4 Kb k 1Kb4',e'6.6 Kb4

Bam Hi Bam Hi Pvull Bam Hi Acc I Nae I Bam Hi Bam Hi Bam Hi

gene5' .....-H3}....ATGAva TGA

T~~gene5'....-tD V.1 VJ2 V,3 V.4CVC. . .3

XMPMP X MA

XFB XMC

FIG. 2. (Upper) Structure of the T4 gene in chromosomal DNA. The V-like (V) and J-like (J), transmembrane (TM), and cytoplasmic (CYT)regions are boxed. The sizes in kilobases of the BamHI restriction fragments are given above. The location offour overlapping X genomic clonesused in mapping and sequence analysis is depicted below. The 5'-untranslated region lies immediately upstream from the ATG initiation codon(L domain); the 3'-untranslated region immediately follows the TGA termination codon (second CYT domain) and continues into the 6.6-kbBamHI fragment. (Lower) Schematic diagram of T4. The domains are designated as L (leader), V (V-like), TM (transmembrane), and CYT(cytoplasmic). J1-i4 immediately follow the corresponding V-like regions. The extracellular cysteine residues that form disulfide linkages areindicated. The position of introns is marked by arrows.

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m a: x a 2o rc = I C:

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FIG. 3. RNA gel blot analysis of RNA derived from mousetissues. Five micrograms of poly(A)+ RNA from various tissuesamples isolated from newborn (NB) and adult (A) mice werefractionated by formaldehyde/agarose gel electrophoresis, trans-ferred to nitrocellulose, and assayed for hybridization to the 32p_labeled pL3T4B cDNA insert described in Fig. 1.

we prepared forebrain and hindbrain RNA from mice span-ning embryonic day 15 through adult (Fig. 4B). In theforebrain, low levels of the 2.2-kb L3T4 RNA are observedfrom embryonic day 15 through postnatal day 9. However,L3T4 RNA levels rise significantly beginning on postnatalday 11 and remain elevated in the adult. In contrast, nohybridization is observed with RNA prepared from hind-brain. We do not know at present whether T4 is expressed byneurons or supporting cells, but as noted the variant mRNAis not likely to be derived from invading hematopoietic cells.Examination of populations of normal peripheral hemato-

poietic cells reveals that T4 is expressed not only in Tlymphocytes but also is expressed in B cells, macrophages,and granulocytes (Fig. 5). These observations are furthersupported by the finding ofT4mRNA in CB (an Epstein-Barrvirus-transformed B-cell line), in U937 (a monocytic cellline), and in HL-60 (a myeloid-granulocyte precursor cell).We note, however, that not all B-cell lines express T4 mRNA

A

28s --

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= : ovIto0 X:

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FIG. 4. (A) RNA gel blot analysis of RNA derived from variousregions of the adult mouse brain. One microgram of poly(A)' RNAfrom thymus, and 5 ,ug of poly(A)+ RNA from whole brain, striatum,cortex, midbrain, hippocampus, cerebellum, medulla, or brainstem,were fractionated, transferred to GeneScreen, and assayed forhybridization to a 32P-labeled pL3T4B RNA probe according toprocedures described by Krumlauf et al. (9). The exposure time ofthe thymus lane was 20% that of the other lanes. (B) Stage-specificexpression of L3T4 in the forebrain. Twenty micrograms of totalRNA was prepared from mouse forebrain at the indicated pre- orpost-natal days and analyzed for L3T4 expression as described inFig. 3. Thearrowheadmarksthe location ofthefull-length 3.2-kbRNA.

FIG. 5. RNA blot analysis of RNA derived from human tissuesand cell lines. Samples include 1 jug ofpoly(A)+ RNA from the T-celllines RPMI (T4-), HSB2 (T4-), CEM (T41), and Jurkat (T4+); fromthe B-cell lines CB (T4+) and Raji (T4-); from the monocytic lineU937 (T4+); from the myleoid-granulocyte precursor HL-60 (T4+);from the human fibroblast line GM (T4-); 5 ,ug ofpoly(A)+ RNA fromhuman cerebral cortex; and 20 ,ug of total RNA from thymocytes,granulocytes, macrophages and E- cells. The analysis was per-formed as described in Fig. 3 using a 32P-labeled pT4B insert (8).

since Raji, an Epstein-Barr virus-transformed B-cell line,expresses no detectable T4 mRNA. In addition, these tissuesand cells were examined for the expression of T8 (or themurine homologue Lyt2) mRNA. T8 is expressed only in thethymus, spleen, and peripheral T cells (data not shown). Thusin contrast to T8, T4 is expressed in a broader range of celltypes than has been suggested from analysis of the T4 proteinon the cell surface.

DISCUSSIONThe organization ofthe T4 gene shows an interesting exampleof exon shuffling over evolutionary time that may provideinsight into the specific function of the T4 molecule and intothe evolutionary origins of the immunoglobulin gene super-family. T4 consists of an extracellular segment comprised ofa leader sequence followed by four tandem VJ-like domains,a transmembrane domain, and a cytoplasmic segment that areeach encoded by separate exons (Fig. 2). The first VJ-likedomain shares significant homology with V regions of im-munoglobulin K chains; the remaining VJ-like domains sharefeatures with both immunoglobulin and T-cell antigen recep-torV and J regions but are most homologous to the polymericimmunoglobulin receptor. Finally, the transmembrane do-main shows considerable homology to the equivalent regionin the P chain of class II MHC molecules. T4, therefore,consists of a collection of exons conserved in several mem-bers of the immunoglobulin gene superfamily that are shuf-fled in different ways to generate a large number of differentmolecules that participate in the immune response.T4 May Reflect a Primitive Immunoglobulin Gene. Data

suggest that T4 interacts with nonpolymorphic regions ofclass II MHC proteins and in this manner facilitates theinteraction of T4+ T lymphocytes with a restricted subset ofantigen-presenting cells bearing class II MHC molecules (22,23). This type of recognition is more reminiscent of primitiveimmune responses that do not appear to involve a diverserepertoire of receptor molecules. T4, although presentlyfunctional in a far more complex immune system, may reflectreceptors operative in more primitive cellular immune re-sponses. Two observations emerge from our analysis of theorganization of the T4 gene that support the suggestion thatT4 may represent a primitive member of the immunoglobulingenefamily. First, the N-terminaldomain ofT4, which sharessignificant homology with the V region of K chains, is split by

- 28S

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LU = 0L1 0 -, 0

- 18S

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an intron. Similar results have been reported for L3T4 (11).Interestingly, introns that interrupt V-like domains have alsobeen observed in the immunoglobulin-like domains of theneuronal cell-adhesion molecule (16), the polymeric immu-noglobulin receptor (K. Mostov, personal communication),and P0, a major protein of peripheral myelin (G. Lemke andR.A., unpublished data). These observations suggest that thecharacteristic 100-amino acid immunoglobulin V domain mayhave evolved from a primordial exon encoding a halfV regionthat later duplicated and fused. Alternatively, the first Vregion of T4 might have evolved by the insertion of intronsequences into a preexisting, complete V region exon. Todate, however, analysis of homologous genes with differingintron-exon organizations have provided evidence for theloss of introns during evolutionary time (24) but have pro-vided no definitive proof for intron acquisition, although thishas been suggested for at least one gene family (25). We favorthe suggestion that the primordial immunoglobulin exonoriginally encoded a half V domain that then duplicated andfused by a recombination mechanism resulting in intron loss.The evolution of the immunoglobulin V domain from theduplication of two half V domains has been suggested fromnucleic acid, protein, and structural analyses (26, 27).A second difference between the structure of T4 and

immunoglobulin genes involves the organization of V and Jsequences. In the immunoglobulin and T-cell antigen recep-tor genes, the V and J regions are encoded by widelyseparated distinct exons and juxtapose only after a somaticDNA recombination event. In the T4 gene, the V- and J-likeregions are adjacent to one another in a single exon. Thus, T4may reflect a primitive gene in which V and J regions werecontiguous prior to the introduction of rearrangement mech-anisms. In this view, the primordial immunoglobulin exonencoded a half V or half VJ domain that duplicated and thenfused with the consequent loss of an intron to generate themore commonly observed 100 amino acid V domain. Con-tiguous VJ regions must have been separated later in evolu-tionary time by the insertion of sequences that facilitatesomatic rearrangement, generating the diversity required ofa more complex immune system.The Function of T4. There are several examples in which

various immunoglobulin domains interact with one another inthe immune system. This property of immunoglobulin genefamily members may be reflected in the association ofT4 withboth class II MHC molecules as well as with the envelopeglycoprotein of HIV. T4, with its polyimmunoglobulin-likeextracellular domain, is thought to interact with class II MHCmolecules on target cell surfaces and, in this manner, directhelper T cells to appropriate antigen-bearing target cells.Class II MHC molecules are also members of the immuno-globulin gene family that most closely resemble immunoglob-ulin constant regions. Interestingly, we have shown that two40 amino acid segments of the HIV exterior envelopeglycoprotein (gp120) also share significant homology withhuman immunoglobulin heavy-chain constant regions, sug-gesting that T4 may recognize immunoglobulin-like struc-tures on both antigen-presenting cells and HIV (7). In supportof this, Lasky et al. (28) have demonstrated that one of theseimmunoglobulin-like regions ofgp120 is essential for efficientbinding of the envelope glycoprotein to T4.

If T4 is indeed a receptor mediating specific cell-cellassociations, then the pattern of expression suggests that itserves such a function in a broader array of cell types than hasbeen suggested. We observe expression of T4 mRNA notonly in T lymphocytes but also in cells of the monocyte-macrophage lineage, B lymphocytes, and granulocytes. More-over, the T4 gene is expressed in a developmentally regulatedmanner in specific regions of the brain. Thus, in contrast to

its counterpart T8, the expression of T4 is not restricted to Tlymphocytes. In T lymphocytes, T4 is thought to play a rolein the recognition of specific target cells. It is tempting tospeculate that T4 plays a more general role in mediating cellrecognition events that are not restricted to the cellularimmune response.

We are grateful to Tom Jessell, Howard Cedar, Tom Maniatis, EdFritsch, Jamie Bigelow, Victoria Stopak, John Fisher, Brian Soda,and Phyllis Jane Kisloff. This work was supported by the HowardHughes Medical Institute (R.A.) and by grants from the NationalInstitutes of Health (R.A. and P.J.M.).

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