A cDNA from <i>Vigna unguiculata</i> encoding the protein kinase p34cdc2

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A cDNA from Vigna unguiculata encoding the protein kinase p34cdc2

KRAUSE, Andréa, EL KOUAISSI, Rachid, BROUGHTON, William John

KRAUSE, Andréa, EL KOUAISSI, Rachid, BROUGHTON, William John. A cDNA from Vigna unguiculata encoding the protein kinase p34cdc2. In: Plant gene register . 1995. p.

Accessionno.X89400

Available at:

http://archive-ouverte.unige.ch/unige:113272

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1 / 1

Plant Gene Register PGR95-076

Krause,A., El Kouaissi,R. and Broughton,W.J. (1995)AcDNAfromVignaunguiculataencoding

the protein kinase p34cdc2 (Accession No. X89400) (PGR95-O76). Plant Physiol.,

iryress toq

:

nz2

^.

A cDNA from Vigna unguiculata encoding the ^protein kinase p34cdc2 (Accession No. ^X8o400 )

AndreaKrause, Rachid El

Kouaissi,

and

William

^J. Broughton

LBMPS, University of

Geneva, 1, ch. de

I'lmperatrice,1292

Chambesy, Switzerland Corresponding author:

AndreaKrause

Fax number: + 4L-22-7320734

E-mail address: krause@sc2a.uni ge.ch

Inoculation of legumes roots

with

rhizobia induces cell division of root cortical cells. Generally cell division depends on the

activity

of the p34cdc7 protein kinase, a key enzyme in the cell cycle (Colasanti et

al.,

1991).

Regulation

of

cdc2

activity

in plants has yet not been

fully

elucidated but appears to ^be similar to other organisms.

In

these organisms enzyme

activity

is regulated at the level of phosphorylation and by the

interaction

with

cyclins

(Doerner,l994;Doonan,

1991). The cdc2- cyclin protein complex is involved in the G2to

M

^phase transition (mitosis)

which

leads to changes associated

with M

phase-specific events (Doerner, L994).ln plants, transcripts encoding the p34cdc2 protein kinase are abundant in meristematic tissue (Martinez

eta\.,7992)

but almost absent

from

tissues composed of terminally differentiated cells (Colasanti

etaI., l99I).

Expression of the cdc2 gene is responsive to treatment

with

external

mitotic

factors such ^as phytohormones

(Hirtetal.,1991;

Martinez eta1.,7992;

Miao etal.,1993)

and Rhizobium

infection (Miao etal-,

L993; Yang

et

al.,

1994). Rhizobial signals

(Nod-

factors) affect the level and

timing

of cdc2 expression (Savoure e/a/., 1994).

A

cdcZhomologous

cDNA

clone

from

Vignaunguiculatawas isolated by screening (at medium hybridization stringency) a root hair

cDNA library

(Krause et

aI.,

1994), using the cdc2-S5

cDNA of

G$cine max (Miao _et aI.,1993) as probe. Only one

positive

clone out of 200,000 recombinant phages was obtained (VuCDC2, GenBank

X8o400).

Sequence analysis showed that VuCDC2 is

of

1309 bp long, including an open reading frame of 882 bp. Further characteristics of the nucleotide sequence are given in Table 1.

A comparison of the V. unguiculatd sequence

with

nucleotide data bases, revealed significant homology to cdc2 genes

of

other plants at the

DNA

and protein levels. Highest homology was found to a

cDNA

clone

encodingthep34cdc2proteinkinaseinV.aconitifulia(Hong etal.,1993).AttheDNAlevel,theoverall

homology wasg'77o. The deduced protein sequence displays ^997o identity

with

only

five

amino acids different. Four ^of, these substitutions are neutral. The amino acid sequence

of

VuCDC2 comprises the

ATP-binding

region, the catalytic domain

for

protein kinase and the PSTAIR

motif,

all which are

highly

conserved

in

p34cdcT protein kinases. Moreover, the functionally important phosphorylation sites are also present in

VuCDC2.The

18 amino acid residues essential for p34cdc2 protein kinase

activity

^are conserved except

for

the substitution

of

the glycine residue at position 184 by a proline which is common in plants.

Together, these data suggest that

VuCDC2

is homologous to an authentic cdc2 gene.

Southern

blot

analysis of genomic

DNA from

V. unguiculatadigested

lvith

various enzymes suggestes that cdc2 homologous genes are present in at least trvo copies in the genome.

Acknowledgments:

We are grateful to DPS Verma

for

making the p34cdc2 protein kinase

cDNA

clone (pcdc2-S5)

of

G. max available. The study was supported by the

University of

Geneva, the Roche Research Foundation and the Fonds National Suisse de la Recherche Scientifique (Project 31-30950.91 and 3 I-40714.94).

DÉPARTEMEN'

O''*O*'NI

^;I"

ËT DE BIOLOGIE VEGETALT

BtBLtorsÈoue

3, Place de I'Université CH.1211 GENÊVE 4

Table L.

Characteristics of the

cDNA

VuCDC2 encoding ap34cdc2 protein _kinase

in

^V. unguiculata Organism:

Vignaunguiculata(L.) Walp., cv

Red Caloona.

Source:

cDNA library

constructed in lambda

NMl149

using

poly(A)+ RNA

isolated from root hairs of V.

unguiculata treated

with Rhizobium

sp. NGR234 (Krause

etal.,

1994).

Clone type and designation:

full-size cDNA

clone named VuCDC2 (GeneBank X84400).

Gene product and function:

Encodes p34cdc2 protein kinase involved in cell cycle control.

Technique of isolation:

Screening of the root hair

cDNA library

under moderate conditions

with

the heterologous probe cdc2-S5

from

G. max

(Miao

et

al.,

L993); sub- cloning into pBluescript

II SK(+)

(Stratagene,

La

Jolla,

CA, USA);

double stranded sequencing

of

exonuclease derived clones using ^the dideoxy-chain

termination

method (Sequenase,

USB,

Cleveland, OH, USA).

Method of

identification:

Sequence homology

with

other cdc2 genes using the

NCBI

Blast network service.

Features of the

cDNA:

The

cDNA

clone VuCDC2 consists

of

1309 bp and possesses an open reading frame of 882 bp. The ORF is

flanked

by untranslated regions of 75 bp at the 5' end and 349 bp at the 3' end. The clone ^has a

poly(A)

tail

of

10 nt.

Features of the predicted protein:

The opèn reading frame encodes ^a polypeptide

of

294amino acids

with

a calculated

Mr

of 34,000 and an estimate d pI of ⁷

.4L

The conserved

ATP-

binding site, the

PSTAIR motif

and the catalytic domain are located in the amino acid sequence at positions 11 to 20, 42 to

57

and 725

to

135, respectively.

In addition,

the phosphorylation sitès T-14,

Y-15

and T-166 as

well

as the amino residues essential

for

p34cdc2 protein kinase

activity

are present.

Literature Cited:

Colasanti J, ^Tyers M, Sundaresan V

^{(1991) Isolation and} characterization of

cDNA

clones encoding ^a functional p34cdc2 homologue

fromZeamays.

Proc Natl Acad Sci

USA 88: 3377-338I

Doerner PW

(1994) Cell cyle regulation

in

plants. Plant

Physiol 106:823-827 Doonan JH

^{( 1991)}

^Cycling

plant cells. Plant J

l:

129-1321-A gricolal

Hirt H, Pay A, Gyorgyey J, ^Bako L, Nemeth K, Bogre L, Schweyen RJ, Heberle-Bors

E,

Dudits D

⁽ 1991) Complementation of a yeast cell cycle mutant by an alfalfa

cDNA

encoding ^a protein kinase homologous to p34cdc2.Proc

Natl

Acad Sci

USA 88:

1636-1640 [Ag.ricolal

Hong Z, Nliao G-H, Verma DPS

⁽¹⁹⁹³⁾ p34cdc2 protein kinase homolog from Mothbean

(Vigna aconitifolia).

Plant

Physiol

101

:

¹³ 99- I

a00 Meclltnçl

[Agri ^{col a]}

Krause A, Sigrist CJA, Dehning I, Sommer H, Broughton WJ [994)Accumulationof

transcripts encoding

alipid

transfer-like proteiÀ during deformation of nbdulation-competent Vigna unguiculataroothairs.

Mol

PlanrMicrob

^I nteract T

:

411 -4 1

8

^lrq gri col al

Martinez MC, Jorgensen J-8, ^Lawton MA, Lamb CJ, Doerner PW

(1992) Spatial pattern of cdcZ expression in relation to meristem

activity

and cell proliferation during plant development. Proc Natl Acad Sci

USA

89 :

736O-73e

^[A gricola]

Miao G-H, Hong Z, Yerma DSP

(1993)

Two

functional soybean genes encoding p34cdc?protein kinases are regulatèd by

different

plant developmental pathways. Proc Natl Acad Sci

USA

9O:943-947 l.a.gricotal

Savoure A, Magyar Z, Pierre M, Brown S, Schultze M, Dudits D, Kondorosi A, Kondorosi

E (1994)

Activatiôn

of the cell cycle machinery and the isoflavonoid biosynthesis pathway by active Rhizobium

meliloti

Nod signal molecules in Medicago microcallus suspensions.

EMBO 13:

1093-1102

Medlinel lAgricola]

Yang W-C, de Blank C, Meskiene I, Hirt H, Bakker J, van Kammen A, Franssen H,

Bisseling T

(L994) Rhizobium Nod factors reactivate ^the cell cycle during infection and nodule primordium formation, but the cycle is only completed in primordium formation. Plant

Cell 6: I4l5-t426 _Medlinel

&

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