JPH07145197A - Ets transcription factor - Google Patents

Abstract

PURPOSE:To obtain a novel ETS transcription factor stimulating the transcription activity of a gene. CONSTITUTION:A site well preserved in the base sequences of an ETS family transcription factor group is used. A cDNA synthesized from a mRNA prepared from an HepG 2 cell is subjected to a PCR reaction to multiply the gene. The base sequence of the produced PCR reactional product is analyzed to give a new ETS transcription factor.

Description

Detailed Description of the Invention

[0001]

FIELD OF THE INVENTION The present invention relates to a novel ETS transcription factor activated by Ras, DNA encoding the ETS transcription factor and an antibody against the ETS transcription factor, and is used in the medical field.

[0002]

The oncogene Ets was found in the retrovirus E26 that causes avian erythroblastosis (Nun.
n, MF, et al., Nature, 306 , 391, 1983). Ets
In the process of cloning protooncogenes, many genes
It was found to have a nucleotide sequence similar to that of ts and is called the Ets family. Ets-1, GABPα, PU1, P
Over a dozen types are known, such as EA-3. These Et
Since the s gene is expected to have a tissue-specific function because the expressed tissues are different, and since it is located near the cleavage site of the chromosomal translocation found in specific human tumors, these Strong attention was paid to the relationship with the pathogenesis of tumors (Rao, VNet al., Sci.
ence, 244 , 66, 1989).

Subsequent studies showed that it has a function of regulating the transcriptional activity of Moroni murine sarcoma virus (Gu
nther, CV et al., Genes Development, 4 , 667, 199
0), it was found that Ets-1 specifically binds to the enhancer region of the α chain gene that constitutes the T lymphocyte antigen receptor and activates transcription (Leiden, JM, Science, 25.
0 , 814, 1990), which attracted attention as a new transcription factor.

The characteristics of the amino acid sequence of Ets family proteins are that, like the myb oncogene product, there is a tryptophan cluster, and this region is called the ETS domain and is known to be a DNA binding site. GGAA / T conserved by IL-2 enhancer, polyomavirus enhancer, HTLV-1, etc. via this domain
Recognize core motifs. Thus, it has been revealed that the Ets family promotes the transcriptional activity of many genes and is deeply involved in the function of cells in response to signals from the outside of cells and oncogene signals such as fms and src. However, there are still many unclear points about the role of these Ets families.

The fibroblast is v-raf or c-H.
Proteins that bind to the Ets binding site of the polyomavirus enhancer, PEA3 element when transformed with a-ras (probably Ets family)
Was known to increase. This protein is observed to increase even in the presence of protein synthesis inhibitors.
It was considered that raf-1 Kinase located downstream of as or phosphorylated by MAP Kinase downstream thereof was directly or indirectly phosphorylated and activated. Since the elucidation of a novel Ets transcription factor opens the way to elucidate the signal transduction mechanism of cells, further elucidation is desired in understanding the growth mechanism of cancer cells.

[0006]

DISCLOSURE OF THE INVENTION The object of the present invention is to provide a novel ETS transcription factor activated by Ras, a DNA encoding the transcription factor and an antibody.

[0007]

SUMMARY OF THE INVENTION In view of the above situation, the present inventors have conducted diligent research to search for a novel ETS transcription factor. First, the structure of the Ets family transcription factor known so far is analyzed, and a well-conserved nucleotide sequence site is selected as a primer. RT-PCR was carried out using a single-stranded cDNA synthesized from mRNA as a template, and the base sequence of the resulting PCR product was subjected to structural analysis, which revealed that known Ets
In addition to the family, a novel ETS transcription factor (hereinafter PEA
3) was confirmed and the present invention was completed. That is, the present invention relates to a novel ETS transcription factor PEA3β containing the amino acid sequence of SEQ ID NO: 1, a DNA containing the DNA encoding PEA3β of SEQ ID NO: 2, and an antibody using PEA3β as an antigen. It is a thing.

[0008] PEA3β of the present invention is Ha-Ras or Ra.
Since the activity is increased by f-1, the above R
It is one of the Ets family that is activated by a downstream factor of as or Ras, and there are three sites that can be substrates of MAP Kinase, in the proline-rich region of the transcription activation site. Among the Ets family transcription factors, there is no factor that is expressed in a wide range of tissues and activated by Ras and Raf, and it was discovered for the first time by the present invention. Elucidation of the growth mechanism of cancer cells, polyomavirus Is important for elucidating the transformation mechanism of

The structure of the present invention is as follows. PEA3
The cDNA encoding β is, for example, (1) mRNA isolated from cells producing an Ets family transcription factor,
(2) A single-stranded cDNA is synthesized from the mRNA, and (3)
From the cDNA structure of the Ets family transcription factor group, a well-conserved portion was selected as a primer, and the above single-chain cD was selected.
Perform RT-PCR using NA as a template. (4) Incorporate the obtained PCR product into an appropriate plasmid, transform the host with it, and (5) conduct DNA analysis of the obtained colony to obtain a novel Ets transcription. The full-length PEA3βcD was selected by selecting the DNA encoding the factor and (6) screening the human cDNA library based on this.
A clone containing NA can be obtained.

As a raw material for preparing mRNA, cells producing an Ets family transcription factor, for example, human hepatocyte mutant strain HepG2 can be used. HepG2
As a method for preparing RNA from guanidine thiocyanate method (Chirgwin JM et al., Biochemistry, 18 , 5
294, 1979) and the like. MRNA can be prepared from the extracted RNA using an oligo (dT) cellulose column. Using the mRNA thus obtained as a template and reverse transcriptase, for example, a single-stranded cDNA is synthesized using the method of Gubler et al. (Gubler, U. at el., Gene, 25 , 263, 1983). . Next, based on the cDNA sequences of the Ets family transcription factors, the well-conserved DN
The A sequence site is selected as a primer, the previously obtained single-stranded cDNA is set, and the gene is amplified by the PCR method.

As the primer to be selected, any part that is well conserved among Ets families can be used, but the DNA binding domain of Ets is well conserved. Among them, it is preferable to prepare a PCR primer based on the nucleotide sequence of a more well-conserved part, and more preferable is 5 ′ primer (corresponding to amino acid QLWQFL) CGGATCCA (A / G) CT (A / G) TGGCA (G / A) TT (T / C) (T / C)
T 3'primer (corresponding to amino acids MNY D / ET / KL / M) GGAATTCA (G / T) C (T / G) T (C / A / G) TC (G / A) TA (G / A)
TTCAT is an example.

A partial library of PCR products can be obtained by incorporating the obtained PCR product into an appropriate vector, for example, pBluescript II, and transforming the host E. coli. As the vector incorporating the PCR product, any vector can be used, such as those derived from Escherichia coli, those derived from Bacillus subtilis, and phages, as long as they are retained in the host cell to replicate and be amplified. As a method for incorporating into a plasmid, for example, Maniatis T et al.
Molecular Cloning, Cold Spring Harbor Laboratory,
Examples include the method described in p239, 1982. The plasmid thus obtained can be introduced into an appropriate host such as Escherichia coli by the calcium chloride method or the calcium chloride / rubidium chloride method (Molecular Cloning, C
old Spring Harbor Laboratory, p249, 1982) and the like.

Since the obtained partial library of PCR products retains the Ets family factor group, for example, by isolating a plasmid from each of the obtained E. coli colonies and analyzing the nucleotide sequence, a known E
Whether it is a ts family factor or a novel factor can be confirmed. Nucleotide sequence analysis was performed by Maxam AM and Gilbert W., Proc. Natl. Acad.
Sci. USA, 74 , 560, 1977) or dideoxy method (M
essing J. et al., Nucl. Acids Res., 9 , 309, 1981
) And the like.

Although the clone having the novel nucleotide sequence does not have the full-length base at this stage, the entire structure can be analyzed by screening a human cDNA library based on this clone. . The human cDNA library is a tissue or cell producing the Ets family transcription factors, for example, human placenta cDNA library, human hepatocyte cell line HepG2cD.
NA library or the like can be used.

Preparation of a HepG2 cDNA library may be carried out according to a conventional method, for example, the above-mentioned method.
MRNA from the single-stranded DN using reverse transcriptase
A, then double-stranded DNA is synthesized. This cDNA is used as a phage vector in λgt11 by the method of Hyunth (DNA Cl
oning, A Practical Approach, 1 , 49, 1985). Then, the HepG2λgt11 library can be obtained by introducing this phage vector into Escherichia coli in which it has been propagated by using the in vitro packaging method.

CDNA is isolated from the clone having the novel Ets transcription factor partial base obtained by the above method,
Using this as a probe, a human cDNA library was subjected to colony hybridization (Gene, 10 , 63, 198).
0) or plaque hybridization (Scienc
e, 196 , 180, 1977) and the like, the desired cDNA clone of the novel Ets transcription factor PEA3β can be searched for. The base sequences of these clones were analyzed by the same method as described above, and if necessary, a part of the cDNA of which the base sequence was determined was synthesized and used as a primer, and the primer extension method (Pr
oc. Natl. Acad. Sci. USA, 76 , 731, 1979) to synthesize a new cDNA, clone a recombinant cDNA by the same method as described above, and analyze the nucleotide sequence to obtain a full-length cDNA of a novel Ets transcription factor. You can confirm the decision. The cDNA sequence of PEA3β of the present invention is shown in SEQ ID NO: 2, and the amino acid sequence is shown in SEQ ID NO: 1.

PEA of the invention consisting of 520 amino acids
The structural features of 3β are as follows. Ets family-Comparison of structure with transcription factor group shows mouse ER8
1, mouse PEA3 and human EF-1 respectively 59
%, 48%, 48% homology. PEA3β
The ETS domain of E. coli has amino acid number 364 in SEQ ID NO: 1.
It corresponds to position -448 and has 96% homology with ER81 and PEA3. Amino acid number 1 in the proline-rich region
It is located at positions 27-269 and is expected to be involved in transcriptional activity together with the acidic region (positions 36-67). Regarding expression of this factor in tissues, the m of 4.0 kb was found in most human tissues.
The expression of RNA was confirmed.

The PEA3β cDNA of the present invention is SEQ ID NO: 2.
The present invention is not limited to the base sequence described, as long as it encodes the amino acid shown in SEQ ID NO: 1, and partial deletion, amino acid exchange, partial addition, etc. are also included in the present invention. It goes without saying that antisense forms based on these base sequences are also included. The same applies to the amino acid sequence.

Novel PEA3β recombinant expression vector and its transformant The PEA3β cDNA obtained by the method described above is incorporated into an appropriate vector, and the vector is transferred into an appropriate host cell to obtain a transformant. .
This can be cultured by a conventional method to produce a large amount of PEA3β from the culture. CDNA encoding PEA3β
A recombinant expression vector can be prepared by recombining A with a known method using a restriction enzyme and DNA ligase downstream of the promoter of a vector suitable for PEA3β expression. Vectors that can be used include Escherichia coli-derived plasmid pBR322, pUC18, and Bacillus subtilis-derived plasmid p.
Examples include UB110, yeast-derived plasmid pSH15, bacteriophage λgt10, λgt11, and SV40, but are not particularly limited as long as they can be replicated and amplified in the host. The promoter and terminator are not particularly limited as long as they correspond to the host used for expressing the nucleotide sequence encoding PEA3β,
Appropriate combinations are possible depending on the host.

The recombinant expression vector obtained in this manner is a recombinant cell method (J. Mol. Biol., 53, 154, 197).
0), protoplast method (Proc. Natl. Acad. Sci. USA,
75, 1929, 1978), calcium phosphate method (Science, 22
1, 551, 1983), in vitro packaging method (Proc.
Natl, Acad. Sci. USA, 72, 581, 1975), virus vector method (Cell, 37, 1053, 1984) and the like are introduced into a host to prepare a transformant. As a host, Escherichia coli, Bacillus subtilis, yeast, animal cells and the like are used, and the obtained transformant is cultured in an appropriate medium according to the host. Culturing is usually carried out at 20 ° C to 45 ° C and pH 5 to 8, and aeration and stirring are carried out if necessary. Separation / purification of PEA3β from the culture may be carried out by appropriately combining known separation / purification methods. Examples of these known methods include salting out, solvent precipitation, dialysis gel filtration, electrophoresis, ion exchange chromatography, affinity chromatography, reverse phase high performance liquid chromatography and the like.

Preparation of Antibodies Polyclonal antibodies for preparing antibodies using PEA3β as an antigen are, for example, subcutaneously, intramuscularly, intraperitoneally, or intravenously inoculated into animals such as mice, guinea pigs and rabbits several times according to a conventional method, and then sufficiently immunized. , Blood collection from such animals, serum separation, anti-P
EA3β antibody is produced. Commercially available adjuvants can also be used. The monoclonal antibody can be produced by a known method. For example, after preparing a hybridoma obtained by cell fusion of spleen cells of a mouse immunized with PEA3β and commercially available mouse myeloma cells, an anti-PEA3β monoclonal antibody can be prepared from the hybridoma culture supernatant or the ascites of the hybridoma-administered mouse. it can. PEA3β used as an antigen does not need to have a whole amino acid structure, and may be a peptide having a partial structure,
It may also be a fusion protein with glutathione transferase and the like. The preparation method may be a biological method or a chemical synthesis method. These antibodies are PEA3β in human biological samples.
It is expected to be used for diagnosis of diseases including cancer. The immunological measurement method of PEA3β may be based on a known method, and can be carried out by any method such as a fluorescent antibody method, a passive agglutination reaction method and an enzyme antibody method.

[0022]

The present invention will be described in detail and specifically with reference to the following examples, but the present invention is not limited to these examples.

Example 1 (1) Preparation of mRNA polyA RNA was prepared from 1.5 × 10 ⁸ HepG2 cells using Fast Track mRNA isolation kit from Invitrogen.
About 10 μg was obtained.

(2) Synthesis of Partial PEA3β cDNA Partial PEA3β cDNA was synthesized by the RT-PCR method. Ets DN in Ets family transcription factors
The A-binding domain was well conserved, and the following primers were synthesized and used based on the nucleotide sequence of the part that was more well conserved. 5'primer (corresponding to amino acid QLWQFL) CGGATCCA (A / G) CT (A / G) TGGCA (G / A) TT (T / C) (T / C)
T 3'primer (corresponding to amino acids MNY D / ET / KL / M) GGAATTCA (G / T) C (T / G) T (C / A / G) TC (G / A) TA (G / A)
TTCAT method is 0.1 μg human liver mRNA and PCR buffer,
dNTPs, DTT and 25 pmol of oligo dT were combined and incubated at 65 ° C. for 5 minutes, then RNAsi
n 40 unit, M-MLV-RT (moloney murine leuke
mia virus-reverse transcriptase ： BRL) 200 unit
Add to make the total volume 25 μl and incubate at 42 ° C for 90 minutes. This reaction was then added to PCR buffer, dNTP
s, 100 pmol of 3'oligonucleotide primer, 10
PCR is performed by further adding 0 pmol of 5'oligonucleotide primer to a total volume of 100 µl. PCR conditions
Aneal temperature 37 ℃ for 30 seconds, extension temperature 72 ℃ for 2 minutes, d
At an enature temperature of 94 ° C., 30 cycles of 1 minute were performed to amplify the DNA and obtain a PCR product. 3 PCR products
Electrophoresed on a% Nu sieve GTG gel, approx. 170-200
A band of bp was cut out, and a part (1/10) thereof was used as a template to perform PCR again.

(3) Integration into pBSKII and transformation The above-mentioned second PCR product was deproteinized with phenol / chloroform, precipitated with ethanol, and then digested with restriction enzymes EcoRI and BamHI. This EcoRI / BamHI fragment was separated on a low melting point agarose and
pBluescriptII SK cleaved with coRI and BamHI and treated with BAP
The (+) vector was ligated with T4 ligase. E. coli XL-1 Blue was transformed with this plasmid to obtain about 300 clones. When the nucleotide sequences of these clones were determined by the dideoxy method (Pharmacia, T7 sequencing kit), clones having a new nucleotide sequence were confirmed in addition to the known Ets family transcription factors GABPα and Ets-2. This base sequence corresponded to positions 1105-1266 of PEA3β cDNA described in SEQ ID NO: 2.

(4) Identification of PEA3β cDNA clone-Production of probe Bam H1 and Hi were cloned into clones having the above novel nucleotide sequences.
It was cleaved with nd III and the DNA fragment was recovered from a 1.2% agarose gel. This DNA fragment was subjected to Klenow enz according to a conventional method.
α- ³² P-dCTP was introduced using yme and used as a probe. -Identification of cDNA clones A filter in which phage DNA was transferred was prepared from the HepG2 cDNA library (about 3.5 x 10 ⁵
The clone) was screened by the DNA hybridization method using the above probe, and two positive clones were obtained. The base sequences of these two cDNAs were determined to confirm the PEA3β total cDNA. PEA3β
The entire cDNA sequence is shown in SEQ ID NO: 2. Escherichia coli XL-1 PEA3β transformed with pBluescript II SK (+) incorporating this cDNA was entrusted to the Institute of Life Science and Technology, Institute of Industrial Science on November 25, 1993 (consignment number FERM P -13982).

Example 2 Expression of cDNA in COS7 cells pcD vector (Okayama, H. and P. Berg., Mol. Cel.
L. Biol., 3, 280, 1983) has a replication origin of SV40 virus and an early promoter, and has a cDNA integrated downstream of this promoter to produce a SV40 T antigen cell line COS7 (Y. Glutzman, Cell. , 23, 175, 1
981), amplification of this recombinant plasmid occurs and transient strong expression of cDNA occurs. Full-Len
excised Bam HI-Xho I fragment containing the entire coding region from pBSPEA3β a clone containing the gth cDNA, pcDV ₁ vector cleaved with Xho I + Hind III (F. Sanger et al., Proc. Natl. Acad. Sci.,
74, 5463, 1977) and DNA ligase to bind to pcDP
EA3β was made. By this operation, the cDNA was incorporated in the correct downstream direction of the promoter. pcDPEA
3β plasmid DNA was prepared and DEAE-dextren
Law (T. Yokota et al., Proc. Natl. Acad. Sci., 82,
68, 1985), the transformed cells transfected into COS7 cells were treated with PEA in the cultured cells after about 1 day.
Produced 3β.

[0028]

The novel ETS transcription factor PEA3β of the present invention
Is a specific factor having the property of being activated by ras. It is useful as a reagent for studying the elucidation of the growth mechanism of cancer cells and the study of the elucidation of the transformation mechanism of poliovirus. Further, the use of an antibody of the factor for the presence / absence of expression in a tissue and for the analysis of a gene having a probe based on the nucleotide sequence of the factor can be applied as a diagnostic agent in a clinical setting. There is expected.

[0029]

[Sequence list]

 SEQ ID NO: 1 Sequence length: 520 Sequence type: Amino acid Topology: Linear Sequence type: Peptide Origin organism name: human Cell type: Liver cell Cell line: Hep G2 Sequence Met Asp Gly Phe Tyr Asp Gln Gln Val Pro Phe Met Val Pro Gly Lys 1 5 10 15 Ser Arg Ser Glu Glu Cys Arg Gly Arg Pro Val Ile Asp Arg Lys Arg 20 25 30 Lys Phe Leu Asp Thr Asp Leu Ala His Asp Ser Glu Glu Leu Phe Gln 35 40 45 Asp Leu Ser Gln Leu Gln Glu Ala Trp Leu Ala Glu Ala Gln Val Pro 50 55 60 Asp Asp Glu Gln Phe Val Pro Asp Phe Gln Ser Asp Asn Leu Val Leu 65 70 75 80 His Ala Pro Pro Pro Thr Lys Ile Lys Arg Glu Leu His Ser Pro Ser 85 90 95 Ser Glu Leu Ser Ser Cys Ser His Glu Gln Ala Phe Gly Ala Asn Tyr 100 105 110 Gly Glu Lys Cys Leu Tyr Asn Tyr Cys Ala Tyr Asp Arg Lys Pro Pro 115 120 125 Ser Gly Phe Lys Pro Leu Thr Pro Pro Thr Thr Pro Leu Ser Pro Thr 130 135 140 His Gln Asn Pro Leu Phe Pro Pro Pro Gln Ala Thr Leu Pro Thr Ser 145 150 155 160 Gly His Ala Pro Ala Ala Gly Pro Val Gln Gly Val Gly Pro Ala Pro 165 170 175 Ala Pro His Ser Leu Pro Glu Pro Gly Pro Gln Gln Gln Thr Phe Ala 180 185 190 Val Pro Arg Pro Pro His Gln Pro Leu Gln Met Pro Lys Met Met Pro 195 200 205 Glu Asn Gln Tyr Pro Ser Glu Gln Arg Phe Gln Arg Gln Leu Ser Glu 210 215 220 Pro Cys His Pro Phe Pro Pro Gln Pro Gly Val Pro Gly Asp Asn Arg 225 230 235 240 Pro Ser Tyr His Arg Gln Met Ser Glu Pro Ile Val Pro Ala Ala Pro 245 250 255 Pro Pro Pro Gln Gly Phe Lys Gln Glu Tyr His Asp Pro Leu Tyr Glu 260 265 270 His Gly Val Pro Gly Met Pro Gly Pro Pro Ala His Gly Phe Gln Ser 275 280 285 Pro Met Gly Ile Lys Gln Glu Pro Arg Asp Tyr Cys Val Asp Ser Glu 290 295 300 Val Pro Asn Cys Gln Ser Ser Tyr Met Arg Gly Gly Tyr Phe Ser Ser 305 310 315 320 Ser His Glu Gly Phe Ser Tyr Glu Lys Asp Pro Arg Leu Tyr Phe Asp 325 330 335 Asp Thr Cys Val Val Pro Glu Arg Leu Glu Gly Lys Val Lys Gln Glu 340 345 350 Pro Thr Met Tyr Arg Glu Gly Pro Pro Tyr Gln Arg Arg Gly Ser Leu 355 360 365 Gln Leu Trp Gln Phe Leu Val Thr Leu Leu Asp Asp Pro Ala Asn Ala 370 375 380 His Phe Ile Ala Trp Thr Gly Arg Gly Met Glu Phe Lys Leu Ile Glu 385 390 395 400 Pro Glu Glu Val Ala Arg Arg Trp Gly Ile Gln Lys Asn Arg Pro Ala 405 410 415 Met Asn Tyr Asp Lys Leu Ser Arg Ser Leu Arg Tyr Tyr Tyr Glu Lys 420 425 430 Gly Ile Met Gln Lys Val Ala Gly Glu Arg Tyr Val Tyr Lys Phe Val 435 440 445 Cys Asp Pro Asp Ala Leu Phe Ser Met Ala Phe Pro Asp Asn Gln Arg 450 455 460 Pro Phe Leu Lys Ala Glu Ser Glu Cys His Leu Ser Glu Glu Asp Thr 465 470 475 480 Leu Pro Leu Thr His Phe Glu Asp Ser Pro Ala Tyr Leu Leu Asp Met 485 490 495 Asp Arg Cys Ser Ser Leu Pro Tyr Ala Glu Val Leu Leu Thr Lys Phe 500 505 510 Leu Ser Gly Gly Val Ala Lys Pro 515 520.

SEQ ID NO: 2 Sequence length: 1800 Sequence type: Nucleic acid Number of strands: Double strand Topology: Linear Sequence type: cDNA to mRNA Origin Biological name: human Cell type: Liver cell line : Hep G2 sequence ATG GAC GGC TTT TAT GAT CAG CAA GTC CCT TTT ATG GTC CCA GGG AAA 48 Met Asp Gly Phe Tyr Asp Gln Gln Val Pro Phe Met Val Pro Gly Lys 1 5 10 15 TCT CGA TCT GAG GAA TGC AGA GGG CGG CCT GTG ATT GAC AGA AAG AGG 96 Ser Arg Ser Glu Glu Cys Arg Gly Arg Pro Val Ile Asp Arg Lys Arg 20 25 30 AAG TTT TTG GAC ACA GAT CTG GCT CAC GAT TCT GAA GAG CTA TTT CAG 144 Lys Phe Leu Asp Thr Asp Leu Ala His Asp Ser Glu Glu Leu Phe Gln 35 40 45 GAT CTC AGT CAA CTT CAA GAG GCT TGG TTA GCT GAA GCA CAA GTT CCT 192 Asp Leu Ser Gln Leu Gln Glu Ala Trp Leu Ala Glu Ala Gln Val Pro 50 55 60 GAT GAT GAA CAG TTT GTC CCA GAT TTT CAG TCT GAT AAC CTG GTG CTT 240 Asp Asp Glu Gln Phe Val Pro Asp Phe Gln Ser Asp Asn Leu Val Leu 65 70 75 8 0 CAT GCC CCA CTC CCA ACC AAG ATC AAA CGG GAG CTG CAC AGC CCC TCC 288 His Ala Pro Pro Pro Thr Lys Ile Lys Arg Glu Leu His Ser Pro Ser 85 90 95 TCT GAG CTG TCG TCT TGT AGC CAT GAG CAG GCT TTT GGT GCT AAC TAT 336 Ser Glu Leu Ser Ser Cys Ser His Glu Gln Ala Phe Gly Ala Asn Tyr 100 105 110 GGA GAA AAG TGC CTC TAC AAC TAT TGT GCC TAT GAT AGG AAC CCT CCC 384 Gly Glu Lys Cys Leu Tyr Asn Tyr Cys Ala Tyr Asp Arg Lys Pro Pro 115 120 125 TCT GGG TTC AAG CCA TTA ACC CCT CCT ACA ACC CCC CTC TCA CCC ACC 432 Ser Gly Phe Lys Pro Leu Thr Pro Pro Thr Thr Pro Leu Ser Pro Thr 130 135 140 CAT CAG AAT CCC CTA TTT CCC CCA CCT CAG GCA ACT CTG CCC ACC TCA 480 His Gln Asn Pro Leu Phe Pro Pro Pro Gln Ala Thr Leu Pro Thr Ser 145 150 155 160 GGG CAT GCC CCT GCA GCT GGC CCA GTT CAA GGT GTG GGC CCC GCC CCC 528 Gly His Ala Pro Ala Ala Gly Pro Val Gln Gly Val Gly Pro Ala Pro 165 170 175 GCC CCC CAT TCG CTT CCA GAG CCT GGA CCA CAG CAG CAA ACA TTT GCG 576 Ala Pro His Ser Leu Pro Glu Pro Gly Pro Gln Gln Gln Thr Phe Ala 1 80 185 190 GTC CCC CGA CCA CCA CAT CAG CCC CTG CAG ATG CCA AAG ATG ATG CCT 624 Val Pro Arg Pro Pro His Gln Pro Leu Gln Met Pro Lys Met Met Pro 195 200 205 GAA AAC CAG TAT CCA TCA GAA CAG AGA TTT CAG AGA CAA CTG TCT GAA 672 Glu Asn Gln Tyr Pro Ser Glu Gln Arg Phe Gln Arg Gln Leu Ser Glu 210 215 220 CCC TGC CAC CCC TTC CCT CCT CAG CCA GGA GTT CCT GGA GAT AAT CGC 720 Pro Cys His Pro Phe Pro Pro Gln Pro Gly Val Pro Gly Asp Asn Arg 225 230 235 240 CCC AGT TAC CAT CGG CAA ATG TCA GAA CCT ATT GTC CCT GCA GCT CCC 768 Pro Ser Tyr His Arg Gln Met Ser Glu Pro Ile Val Pro Ala Ala Pro 245 250 255 CCG CCC CCT CAG GGA TTC AAA CAA GAA TAC CAT GAC CCA CTC TAT GAA 816 Pro Pro Pro Gln Gly Phe Lys Gln Glu Tyr His Asp Pro Leu Tyr Glu 260 265 270 CAT GGG GTC CCG GGC ATG CCA GGG CCC CCA GCA CAC GGG TTC CAG TCA 864 His Gly Val Pro Gly Met Pro Gly Pro Pro Ala His Gly Phe Gln Ser 275 280 285 CCA ATG GGA ATC AAG CAG GAG CCT CGG GAT TAC TGC GTC GAT TCA GAA 912 Pro Met Gly Ile Lys Gln Glu Pro Arg Asp Tyr Cys Val A sp Ser Glu 290 295 300 GTG CCT AAC TGC CAG TCA TCC TAC ATG AGA GGG GGT TAT TTC TCC AGC 960 Val Pro Asn Cys Gln Ser Ser Tyr Met Arg Gly Gly Tyr Phe Ser Ser 305 310 315 320 AGC CAT GAA GGT TTT TCA TAT GAA AAA GAT CCC CGA TTA TAC TTT GAC 1008 Ser His Glu Gly Phe Ser Tyr Glu Lys Asp Pro Arg Leu Tyr Phe Asp 325 330 335 GAC ACT TGT GTT GTG CCT GAG AGA CTG GAA GGC AAA GTC AAA CAG GAG 1056 Asp Thr Cys Val Val Pro Glu Arg Leu Glu Gly Lys Val Lys Gln Glu 340 345 350 CCT ACC ATG TAT CGA GAG GGG CCC CCT TAC CAG AGG CGA GGT TCC CTT 1104 Pro Thr Met Tyr Arg Glu Gly Pro Pro Tyr Gln Arg Arg Gly Ser Leu 355 360 365 CAG CTG TGG CAG TTC CTG GTC ACC CTT CTT GAT GAC CCA GCC AAT GCC 1152 Gln Leu Trp Gln Phe Leu Val Thr Leu Leu Asp Asp Pro Ala Asn Ala 370 375 380 CAC TTC ATT GCC TGG ACA GGT CGA GGC ATG GAG TTC AAG CTG ATA GAA 1200 His Phe Ile Ala Trp Thr Gly Arg Gly Met Glu Phe Lys Leu Ile Glu 385 390 395 400 CCG GAA GAG GTT GCT CGG CGC TGG GGC ATC CAG AAG AAC CGG CCA GCC 1248 Pro Glu Glu Val Ala Arg Arg Trp Gly Ile Gln Lys Asn Arg Pro Ala 405 410 415 ATG AAC TAT GAC AAG CTG AGC CGC TCT CTC CGC TAT TAC TAT GAA AAG 1296 Met Asn Tyr Asp Lys Leu Ser Arg Ser Leu Arg Tyr Tyr Tyr Glu Lys 420 425 430 GGC ATC ATG CAG AAG GTG GCT GGA GAG CGA TAC GTC TAC AAA TTT GTC 1344 Gly Ile Met Gln Lys Val Ala Gly Glu Arg Tyr Val Tyr Lys Phe Val 435 440 445 TGT GAC CCA GAT GCC CTC TTC TCC ATG GCT TTC CCG GAT AAC CAG CGT 1392 Cys Asp Pro Asp Ala Leu Phe Ser Met Ala Phe Pro Asp Asn Gln Arg 450 455 460 CCG TTC CTG AAG GCA GAG TCC GAG TGC CAC CTC AGC GAG GAG GAC ACC 1440 Pro Phe Leu Lys Ala Glu Ser Glu Cys His Leu Ser Glu Glu Asp Thr 465 470 475 480 CTG CCG CTG ACC CAC TTT GAA GAC AGC CCC GCT TAC CTC CTG GAC ATG 1488 Leu Pro Leu Thr His Phe Glu Asp Ser Pro Ala Tyr Leu Leu Asp Met 485 490 495 GAC CGC TGC ACC AGC CTC CCC TAT GCC GAA GTT TTG CTT ACT AAG TTT 1536 Asp Arg Cys Ser Ser Leu Pro Tyr Ala Glu Val Leu Leu Thr Lys Phe 500 505 510 CTG AGT GGC GGA GTG GCC AAA CCC TAGAGCTAGC AGTTCCCATT CAGGCAAACA 1590 Leu Ser Gly Gly Val Ala Lys Pro 515 520 AGGGCAGTGG TTTTGTTTGT GTTTTTGGTT GTTCCTAAAG CTTGCCCTTT GAGTATTATC 1650 TGGAGAACCC AAGCTGTCTCTCGGAGAG 1710 TGGGAACAGG GAGGGGCAGA 1
00

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Claims (3)

[Claims] 1. An ETS transcription factor comprising the amino acid sequence represented by SEQ ID NO: 1. 2. An ETS containing the nucleotide sequence of SEQ ID NO: 2 which codes for the ETS transcription factor of claim 1.
Transcription factor cDNA 3. An antibody using the ETS transcription factor according to claim 1 or a partial peptide thereof as an antigen.