Geraldine Butler a,, a Department of Biochemistry, Universit 3

b Department of Genetics, University o]

Received

tact

Saccharomyces cerevisiae gene (HTN1) that encodes a ho ~A) was identified, cloned and sequenced. The two proteins GSP1, GSP2, and PRP20 proteins in an intracellular signall

~ay. RanBP1 homologues also exist in worms and rice.

9rds: Ran/TC4 protein; RCC1; GTP binding protein; RanBP1; HTF!

L mammalian cells the product of the R C C 1 gene is lved in the regulation of progression through the cell ~_ and in narticular the control of chromosome conden-

gnanlng pamway equivalent to me 1

HTF9A

implicated in regulation of chromosomes, progn

.ntrol of chromosome conden- and export, and regulal rying a temperature sensitive cently, it has been sugg 9sis if the cells are shifted to role in transport of protq luring S phase, even though [6-8]. plete, suggesting that RCC1 While searching for

determining the cell cycle R a n / T C 4 , Coutavas et on [1,2]. Analysis of proteins GTPase activating prote :o the identification of a GTP binding protein (RanBP /TC4 , which is a member of product of a previously pecifically catalyses the ex- [10]. RanBP1 and Ran// ' has been suggested that the their interaction is othe ,'r with other proteins), acts as RanBP1 function smission pathway that detects R C C 1 / R a n control of o equivalent signalling system Using the program T t Saccharomyces cerevisiae, larity between mouse R homologue encoded by the upstream of the neutral n//TC4-1ike genes, GSP1 and centromere of yeast chr tial gene that is highly tran- this region by polymel :xpressed at low levels in a nucleotides with nner, and is not required for T A G C G A A G A T A A //PRP20 proteins have been GACGCAAGTAAC-3 '

K eyw o r ds :

In involved cycle, and in particular the control of sation [1]. Hamster cells carr' mutation in R C C 1 enter mitosis the restrictive temperature durin DNA synthesis is not complete, plays an important role in monitoring of DNA replication that interact with RCC1 led to binding nuclear protein, Ran/ the ras family [3]. RCC1 specifica change of GTP on Ran. It R C C 1 / R a n complex (together a component of a signal transmission unreplicated DNA [1]. An seems to exist in the yeast which contains an RCC1 P R P 2 0 gene [4], and two Ran G S P 2 [5]. GSP1 is an essential scribed, whereas G S P 2 is ex carbon-source dependent manner cell viability [5]. The RCC1

~The nucleotide sequence data reported submitted to the EMBL/GenBank Data Lib number Z33503

Biochimica et Biophysica Acta 1219 (1994

Short Sequence-l?

logue of mammalian

a,* Kennc University College Dl

~ Dublin, Tri

27 May 199

homologue c are 51% i~

gnalling pathw

of chromosome

fically

that ion

e t Bio i

lding protein 1

~lfe b

9ublin 4, Ireland ublin 2, Ireland

binding protein 1 (RanBP1, tuence. The HTN1 protein m to the mammalian RanBP1-F

of DNA replication ress into mitosis, m

ulation of gene expre ggested that R a n / T C

9roteins across the nucl

mouse proteins thai al. [9] identified

notein (Ran-GAP), as , RanBP1). RanBP1 was fo

characterised mouse ~TC4 bind tightly bu erwise unknown. It

unctions as an interme cell-cycle progressior TBLASTN [11] we il RanBP1 and yeast D

trehalase gene (N'. chromosome IV [12].

,merase chain reaction sequence 5 ' - G G G A A

~ T A A G - 3 ' and 5'-,t and genomic DNA

e strain H9. The 0.6 kb PC I vector (Invitrogen) and sequenced using an Appli sequencer. Our sequence

deletions as compared to tl

agacgaagaagttctttacaaggtcagagccaagcttttcagattc

) A D A K E W K E R Ltgccgatgccaaggaatggaaagaaagaggtactggtgactgtaag

' L K N K K T N K V R cttgaagaacaaaaagactaacaaggttagaatattgatgagaaga

) K T L K I C A N H ~caagaccttaaagatttgtgctaaccacatcattgctccagaatac

' L K P N V G S D R tttgaagcctaacgttggttctgatagatcttgggtgtatgcttgt

A D I A E G E A E

agcagatattgcagaaggtgaagcagaagccttcacttttgctatc

~ F G S K E N A D K latttggcagtaaggaaaatgctgataaatttaaagaagaatttgaa

[ A Q E I N K K A

agctcaagaaatcaacaaaaaggcttag

1. Nucleotide and deduced amino acid sequen~ of the Saccha-

12], resulting in an open reading ~ame of 201 codons, tming that translation begins at the first available me- nine codon (Fig. 1). The gene, designated HTN1, is ,ted 568 bp upstream of NTH1 in a head-to-head ngement. Codon usage in HTN1 is quite stronNy

cell cycle by regulating acids across the nuclear system will be facilitate tion of Saccharomyces

References

Dasso, M. (1993) Trend,, Nishimoto, T., Eilen, E.

l d e x - 0.49; Ref. [13]), which Bischoff, R.F. and Ponst quite highly expressed. The Aebi, M., Clark, M.W., highly charged (42% of the Mol. Gen. Genet. 224, 7

approximately equal numbers Belhumeur, P., Lee, A., M.W. (1993) Mol. Cell. Moore, M. S. and Blobe

1 and mouse RanBP1 proteins Goldfarb, D.S. (1994) C The binding protein is much Melchior, F., Paschal, 13

n is the GTPase itself (82% Biol. 123, 1649-1659. n and yeast GSP1) but more Coutavas, E., Ren, M., ( nucleotide exchange proteins M.G. (1993) Nature 366

Bressan, A., Somma, M 'RP20 proteins are only 33% N.G., Gilbert, D.J., Jenl hes of the dbEST database of 201-209.

Altschul, S.F., Gish, W.

S in the nematode Caenorhab- (1990) J. Mol. Biol. 215 bsperm Oryza sativa (Fig. 2). [12] Kopp M., Miiller n. al

4766-4774. l sequences in animals, plants, [13] Lloyd, A.T. and Sharp,

c o m m o n signalling pathway [14] Boguski, M.S., Lowe, 1 in (RanBP1 or HTN1), a GT- Genet. 4, 332-333.

Fig. romyces cerevisiae HTN1 gene.

al. [12 assumm thionine located arran biased (Codon Adaptation Index suggests that the gene is predicted HTN1 protein is residues are charged), with of acidic and basic residues.

The predicted yeast HTN1 are 51% identical (Fig. 2). more poorly conserved than identity between human Ran slowly evolving than the (human RCC1 and yeast PRP20 identical). TBLASTN searches single-pass cDNA sequences HTN1/RanBP1 homologues ditis elegans and the angios[ The conservation of RanBP1 and fungi suggests that a comprising a binding protein

Butler, K.H. Wolfe/Biochimica et Biophysic

D K K E E A C. e~ ~acaagaaggaagaggct 48 mous~

yeast

S M F G G K =ccatgt t t ggtggtaag 96 C. e:

maus~ E E D T K E yeast ~aagaagataccaaggag 144

rice E S P D I H C.e.

~aatcaccagatatccat 192 mous( yeast

D V K T M E

~atgt taagacaatggaa 240 rice

c. e. A K L F R F mous(

288 yeast

G T G D C K 336 mous~

yeas~

I L M R R

384 Fig. 2 Tilde.,

I I A P E Y 4 3 2 Caent

rived S W V Y A C

4 8 o lower mous(

A F T F A I 528

F K E E F E Pase 5 v 6 chan

otes. 6O6

[1] [2] [3] [4]

[51

[61 [71 [81

[91

[10]

[14] also identified [11]

.... dttln M

~DKKPWDKKEEAAPKPPS SAVF SMFGGKKA

~eyePe---veFkPviplpdlvevKTgEEDE ~NHDPQ ..... FEPIVSVPE-QEIKTLEEDE ~DDAPES PD IHFEPV~LEK-VDVKTMEEDE

.... KetgkvRlvMRqaKY cEnKERG iGDiKiLKsnd-nkyRivMRReclv ? EWKERGTGDVKLLKHKEKGT I RLLMRRDKT - EWKERGTGDCKFLKNKKTNKVR I LMRRDKT

3SDkscVxa- rAgl r r relkeem fAI RFgsv %ipnvlt fmcpgllrr .... ]SDRAWVWNTHADFADEC PKPELLAI RFLN~ ~SDRSWVYACTAD IAEGEAEAFTFAI RFGSg

REKKG PGKNDNAEKVAEKL EAL SVR EARE E~ INKKA

predicted protein sequences of Ra ms that are identical between yea ms and rice sequences are incon NA sequencing runs; these seque xcept for positions that are ider

or GSP1/GSP2) and a EC1 or PRP20) may exist i lays may control progressi

the transport of prot tclear membrane. Further

tcilitated by the ease of gel cerevisiae.

Trends Biochem. Sci. 18, 96-1 and Basilico, C. (1978)

Ponstigl, H. (1991) Nature 35 Vijayraghavan, U. and

72-80. Tam, R., DiPaolo, T., Fc Biol. 13, 2152-2161.

Blobel, G. (1993) Nature 365, Curr. Biol. 4, 57-60. B., Evans, J. and Gerace,

Oppenheim, J.D., D'Eust." ,585-587 .

M.P., Lewis, J., Santolam~ Jenkins, N.A. and Lavia, P.

W., Miller, W., Myers, E.'W :, 403-410.

and Holzer, H. (1993) J.

P.M. (1992) J. Hered. 82 T.M.J. and Tolstoshev, C