Complementary DNA Cloning and Chromosomal Mapping of a ... protein traffic.1-2 The superfamily consists of three ma- ..... human chromosome 1 (Fig. 3 (a)).
DXA
RESEARCH
4. 301 305 (1997)
Short Communication
Complementary DNA Cloning and Chromosomal Mapping of a Novel Phosphatidylinositol Kinase Gene Tosliiyuki
SAITO.1*
Tada-aki HORI
Xaohiko
SEKI. 2
Hideshi ISHii.1 Miki
OHIRA. 2
Akiko
HAYASHI. 1
Sumie
KOZUMA. 1 and
1
Genome Research Group. National Institute of Radiological Sciences. 4-9-1 Anagawa. Inage-ku. Chiba 263. Japan1 and Laboratory of Gene Structure I. Kazusa DNA Research Institute. 1532-3 Yana. Kisarazu. Chiba 202. Japan2 (Received 5 August 1997) Abstract
It was recently revealed that an expanding superfamily of lipid/protein kinases with a phosphatidylinositol (PI) kinase domain regulates various cellular processes such as signal transduction. cell cycle and intracellular protein traffic.1-2 The superfamily consists of three major families. The first is the PI3 kinase family, which catalyzes the phosphorylation of PI at the D3 position of the inositol ring. The PI3 kinases have been found to associate with the receptors of growth factors having tyrosine kinase activity'5 and they are believed to play a role in the transduction of signals triggered by a variety of growth factors and hormones. The second is the PI4 kinase family, which catalyzes the phosphorylation of the D4 position of PI and is considered to be involved in an important step of intracellular signalling. The third is the ATM family, some members of which have been demonstrated to be protein kinases although their authentic substrates have not yet been identified. This family includes FRAP. FRP1. DNA-PKcs and ATM proteins that are involved in cell cycle regulation, checkpoint control and maintenance of genome integrity.4 9 The still increasing number of superfamily members and the conserved amino acid stretches shared among them encouraged us to search for other members by utilizing a degenerated oligonucleotide-primed polymerase chain reaction (PCR) technique described *
Communicated by Mituru Takanami To whom correspondence should be addressed. Tel. 81-43-2063151. Fax. 81-13-251-9818. E-mail: t-saitoSnirs.go.jp
previously.10 To identify unknown superfamily members for better comprehension of the cell cycle regulation and signal transduction systems, we carried out a reverse transcriptase PCR (RT-PCR) experiment with degenerative primers (S'-G-A-T/C-G-A-T/C-C-T-N-C/A-GN-C-A-A/G-G-A-3' and 5'-N-C-C-A/G-A-A-A/G-T-CA/T/G-A-T-A/G-T-G-3') for the conserved amino acid sequences (D/E-D-L-R-Q-D/E and H-I-D-F-G) of the superfamily and a variety of mRNA sources. Cloning and sequencing of a fraction of the RT-PCR products suggested the existence of at least two novel members. One is another member of PI3 kinase family, the other is the new PI4 kinase described in the present study. The nucleotide sequence of the 3.5-kb cDNA clone and predicted amino acid sequence are shown in Fig. 1. The nucleotide sequence data reported here will appear in the DDB.I. EMBL. and GenBank nucleotide sequence databases under accession number AB005910. The predicted protein of 961 amino acids has a calculated molecular weight of approximately 107 kDa. The overall structure of the protein showed extensive similarity with the previously reported PI4 kinase. 11 implying that the protein described in this paper is a PI4 kinase. Very recently, another PI4 kinase member called PI4 kinase 3 was reported in humans and in the rat. 1 2 1 3 which was sensitive to wortmannin and has the even more striking similarity to our present sequence. Although our cDNA clone was isolated completely independently of the above report, our sequence and that of PI4 kinase 3 seem to be
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A cDXA for a putative new member for phosphatidylinositol kinase family was cloned from an adult human whole brain cDNA library. The predicted translation product was composed of 961 amino acid residues and contained a sequence feature characteristic for lipid/protein kinases. The messenger RNA was ubiquitously expressed in various tissues, while relatively higher expression was observed in heart, skeletal muscle and testis. The chromosomal location of the gene was determined by fluorescence in situ hybridization and PCR-based analyses with both a human/rodent monochromosomal hybrid cell panel and a radiation hybrid mapping panel. Key words: PI4 kinase: RT-PCR: Iq21
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70 80 110 90 120 100 GGAAATTGAATGAGATTCTTGGAAGCTCGAAGTCTGGCTGTGGCCATGGGAGATACAGTA * M R F L E A R S L A V A M G D T V 130 140 170 150 180 160 GTGGAGCCTGCCCCCTTGAAGCCAACTTCTGAGCCCACTTCTGGCCCACCAGGGAATAAT V t P A P L K P T S E P T S G P P G N N 190 200 230 210 240 220 GGGGGGTCCCTGCTAAGTGTCATGACGGAGGGGGTCGGGGAACTATCAGTGATTGACCCT G G S 1 1 S V J T E G V G E L S V I D P 250 260 290 270 300 280 GAGGTGGCCCAGAAGGCCTGCCAGGAGGTGTTGGAGAAAGTCAAGCTTTTGCATGGAGGC E V A Q K A C Q E V L E K V K 1 L H G G 310 320 350 330 360 340 GTGGCAGTCTCTAGGAGAGGCACCCCACTGGAGTTGGTCAATGGGGATGGTGTGGACAGT V A V S S R G T P L E L V N G D G V D S 370 380 410 390 420 400 GAGATGCGTTGCCTAGATGATCCACCTGCCCAGATCAGGGAGGAGGAAGATGAGATGGGG F I R C 1 D D P P A Q I R E E E D E M G 430 440 470 450 480 460 GCCGGTGTGGCCTCAGGCACAGCCAAAGGAGCAAGAAGACGGCGGCAGAACAACTCAGCT A A V A S G T A K G A R R R R Q N N S A 490 500 530 510 540 520 AAACAGTCTTGGCTGCTGAGGCTGTTTGAGTCAAAACTGTTTGACATCTCCATGGCCATT K Q S W L L R L F E S K L F D I S M A I 550 560 590 570 600 580 TCATACCTGTATAACTCCAAGGAGCCTGGAGTACAAGCCTACATTGGCAACCGGCTCTTG S Y 1 Y N S K E P G V Q A Y I G N R E F 610 620 650 630 660 640 TGCTTTCGCAACGAGGACGTGGACTTCTATCTGCCGCAGTTGCTTAACATGTACATCCAG C F R N E D V D F Y L P Q L L N M Y I H 670 680 710 690 720 700 ATGGATGAGGACGTGGGTGATGCCATTAAGCCCTAGATAGTCCACCGTTGCCGCCAGAGC M D E D V G D A I K P Y I V H R C R Q S 730 780 740 77
