Jpn. J. Infect. Dis., 53, 2000

Laboratory and Epidemiology Commumications
Fv‑ 1 Restriction of Murine Leukemia Ⅵms May Not Necessarily

Be at Cytoplasmic‑Nuclear Transport Phase
Takashi Odawara*

AIDS Resewch Center, National Institute ofInfectious Diseases,
Gakuen 4‑ 7‑1, Musashimurayama‑shi, Tob70 208‑0011
Communicated by Hiroshi Yoshikura(Accepted May 29, 2000)

Fv‑ 1 resbiction ofmurine leukemia virus (MuIN) determinedby two amino acids in p30 (CA) of the viral protein (I‑2) isIn order to examine the formation of linear and circularproviruses in the infected cells, cells plated in an amount ofconsidered to occur during the post‑penetration phase of

2 XIO6 cells/10 cm dish were infected withthe viruses in theinfection (3‑5). Since the formation of circular proviral DNA,

presence of 0.8 LLg/ml of polybrene for 5 h. At indicatedwhich is considered to reflect the entrance of pre‑integration

periods a洗er the start of infection, DNAs were extracted ac‑complex into the nucleus has been seen to be impaired incording to Hirt's method (ll). One血ird or the DNAs

the restrictive cells (6‑7), it has been speculated that theextracted from each plate was electrophoresed in 0.8% agarose

cytoplasmic‑nuclear transport of the provlruS Was inhibitedgel, and analyzed by Southem blotting uslng an Xbal‑Xbal

in restrictive cells. However, whether Fv‑ 1 acts in cytoplasm,(#57661#8 1 13) fragment of Moloney MuLV env as a probe･As shown in Figure 3‑A,血ough fbmation of血e clrCularin the nucleus, or in bo血is still unknown.

I report here that an MuLV, which was constructed byreplacing the gag region Of NB‑tropic Moloney MuLV with
N‑tropic WN 1 802N‑derived gag, N‑Mop (8), did not showproviral DNA ofWN 1 802N was inhibitedinC5 7BL/6‑derived
B‑type YH‑7 cclls, that ofN‑MoF was not. That ofB‑MoFdA
in N‑typeNIH3T3 cells was inhibited as expected (Fig. 3l

any inhibition offormation of the circularintermediate, though

B).
The above observation showed that the Fv‑1 restriction
could take place without inhibition of the formation of circularDNA. The circular formis not considered a pre‑integration

its titration pattem was that of atypical N‑tropic virus.
Figure 1 shows the structures ofN‑MoF and a similar B‑
tropic constmct, B‑MoFdA (9). In XC assay (10), both N‑
MoF and B‑MoFdA showed the expected titration pattem ofmolecule itself but rather a by‑product (12). Therefore, thisdiscrepancy may not be surpnslng. However, as the circular

N‑tropic and B‑tropic viruses (Fig.2).

(gagxpo/xenv)

SallPvu I

WN1 802N

N M0 ド一●■1●‑■1‑lももミMo･48
A鉱ⅠⅠ: PvuI:Sal I(#363日榊19)

BN｢FdA.iWP…L l:(#3 705)｢l1‑‑■■■…｣''1‑■‥忘]

Illl

I

WN1 802B
AaHISatIFig. 1. Structure ofrecombinant viruses (See refs. Band 9).
Thick丘lled portion is derived &om WN1802N,thick shaded portion丘om WN1 802B,and thin portion舟om Moloney MuLV Mo48.

*Corresponding author: E‑mail: odawara@cshl.org

88

Jpn. J. Infect. Dis., 53, 2000

N‑Mop

B･MoFdAl､.
1､□ : NIH3T3◆Il

10 0 0 1""

B蕋○0 :YH‑7l､.

"l一､..

1IIト,0 :SC‑1lI

8 卜､lllー■‑⊂〉⊂〉⊂〉一･l‑WN1⊂〉⊂>⊂I⊂)⊂>〇O0一一OOO0■‑⊂〉802Nr

2WN1802B⊂〉⊂Ir2ヽ ヽ ヽ ヽ ヽ ヽ a ヽ ヽヽ咾､食.
ヽVヽヽ ､､J丁白J. lll
一一⊂〉O■一∴lllllll■⊂〉〇▼･･⊂〉く⊃⊂〉▼‑Fig. 2. Titration ofN‑Mop, B‑MoFdA, and WN1802N/B on N‑type NIH3T3, B‑type YH‑7, and Fv‑I
unrestrictive SC̲ I cells.

The linear curve fitting was done by CA‑Cricket Graph III.

Vertical axis: number ofplaquesper plate.
Horizontalaxis: dilution of viruses.

A WN1802N N･MoF Mo･48onbonbonformis strictly nuclear (13), the restriction ofN‑MoF wasexerted probably after entrance to the nucleus; i.e., it wasb

inferred that Fv‑ I restriction can occur in the nuclear phase.

As WN1802N and N‑MoF had gag reg10nS With the same
sequence, it is speculated that a region Other than gag is able

to determine the impaired formation of circular DNA.

REFERENCES

1. DesGroseillers, L and Jolicoeur, P. (1983): Physicalmapplng Of the Fv‑1 tropism host range determinant ofBALB/c murine leukemia viruses. J. Viro1., 48, 685‑696.

!一印欝■‑

2. Ou, C. Y., Boone, L R, Koh, C. K., Tennant, R. W. andYang, W. K.(1983): Nucleotide sequences of gag‑pol
regions that detemine the Fv‑ I host range propertyof

蓋u書:13尊t :･三二B

M0‑48

o

N･MoF

nbon

BALB/c N‑tropicand B‑tropic murine leukemia viruses.
J. Viro1., 48, 779‑784.
3. Yoshikura, H. (1973): Host range conversion of the murine

sarcoma‑leukemia complex. J. Gen. Viro1., 19, 321‑327.
4. Huang, A. S., Besmer, P., Chu, L. and Baltimore, D.

B‑MoFdAbon(1973): Growth of pseudotypes of vesi.cular stomatitis
virus with N‑tropic murine leukemia vlruS coats in thecells resistant to N‑tropic viruses. J. Viro1., 12, 6591662.
5. KrontiriS, T., Soeiro, R. and Fields, B. N. (1973): Hostb

resbiction offriend leukemia virus. Role of the viraloutercoat. Proc. Natl. Ac礼d. S°i. USA, 70, 2549‑2553.

6. Jolicoeur, P. and Rassart, E. (1980): Effect ofFv‑1 geneproduct on synthesis of lineafand supercoiled viral DNAin cells infected with murine leukemia virus. J. Viro1.,
33, 183‑195.

Fig. 3. Formation of extrachromosomalproviralDNA a触r infection.

Eachvirus stock of comparable titer on Sc‑I cells･was infected to

7. Yang, W. K., Kiggans, ∫. 0., Yang, D. M., Ou, C. Y,

SC‑1 (o), NIH3T3 (n), or YH‑7 (b) cells. Atthc indicated hours a鮎r

Termant, R. W., Brown, A. and Bassin, R.H. (1980):
Symthesisand circularization ofN‑and B‑tropic retroviral
DNA Fv‑ 1permissive and restrictive mouse cells. Proc.

infection, extrachromosomalDNAswere extracted according to Hirt's
method (I I). The DNAswere electrophoresed in 0.8% agarose gel,and analyzed by Southern‑ blotting using Moloney‑MuLV env probe.
L: linear form. cc: closed circular form.

Natl. Ac礼d. S°i. USA, 77, 2994‑2998.

8. Kawana, A., Iwamoto, A., Odawara, T. and Yoshikura,
H. (1997): Host range conversion ofmurine leukemia
virus resulting from recombination with endogenous

89Jpn. J. Infect. Dis., 53, 2000

vims. Arch. Viro1., 142, 139‑149.
9. Doi, K., Kawana, A., Iwamoto, A., Yoshikura, H. andOdawara, T･ (1997): One base change iP sufficient for

369.

12･ Fujiwar.a, T･ and Mizuuchi, K･ (1988): Retroviral DNA
integrat10n: Structure Ofanintegration intermediate. Cell,

host range conversion of murine leukemla Virus from B

54, 497‑504.

10. Rowe, W. P., Pugh, W. E.and Hartley, J. W. (1970): Plaque

13. Guntaka, R. V, Richards, 0. C., Shank, P. R., Rung, H.
J. and Davidson, N. (1976): Covalently closed circular

assay techniques for murine leukemia viruses.Virology,DNA of aviansarcoma virus: purification from nuclei ofto NB tropism. Arch. Viro1., 142, 1889‑1894.

42, 1136‑1139.

infected quail tumor cells and measurement by electron

microscopy and gel electrophoresis. J. Mol. Bio1., 106,

1 I. Hirt, B. J. (1967): Selective extraction ofpolyoma DNA

from infected mouse cell cultures. J. Mol. Bio1., 26, 3651

337‑357.

Laboratory and Epidemiology Commumications
SuⅣeillance of Poliovims‑Isolates in Japan, 1 999

Tetsuo Yoneyama* , Yoshiaki Karojil , Kanako Watanabe2, Misako Tsuchiya3,MamoruNakano4 and Tatsuo Miyamura
Department of Viy10logy II, National Institute ofInfectious Diseases,
Musashimurの,ama 208‑0011,

JKyoto CiOJ Institute ofHea/th and Environmental Sciences,
Kyoto 604‑8845,

2Niigata Pylefectural Resewch Laboratopyfor Health and Enviy10nment,
Niigata 950‑21 44,

3Fukushima Institute of Health and Environmental Sciences,
Fukushima 960‑81 63 and4Nara Prefectural Institute ofPublic Health,Nara 630‑8131
Commumicated by Tatsuo Miyamura(Accepted May 29, 2000)

In 1999, I I polioviruses were isolated from nine clinical

Asshown in Table I, all of the examined polioviruses werevaccine‑derived strains. Type3 poliovirus was isolated frompatientsinfourprefectures. Samples of the viruses were sent toNational Institute of Infectious Diseases (NIID)and subjectedto the intratypic differentiation by PCR‑restriction fragment

the stool specimen of a poliomyelitis patient in Kyoto, andlength polymorphism method developed by Radu Crainic ( 1 ).

of the paralysisinthe Kyoto case occurred in November 1 998,

examined forintratypic differentiationin1999. Sincethe onset

Table I. Characterization ofpoliovirus isolates in 1999

Date or vaccination

CodeAgeSexin the patient in the area.Dnase.of Paamb,Fi:g dFilignicoaSlis Serotype

諾監禁ati｡n99‑kyot0‑ 12YM

98‑1 0‑28(2nd)98‑ll‑19

98‑1ト19

Poliomyeliti s

Polio̲3Vacc ine‑like

9 9 ‑kyot0‑27M

F

98‑4‑16

98‑5‑2

98‑5‑2

Aseptic meningiti s

Polio̲2

Vaccine‑ like

99‑Niigata‑ 1FM
M

98‑1 0‑239914‑16

99‑4‑599 ‑4‑9Gastroenteritis

Polio̲ 1 &2

ⅥlCCine‑ like

98‑1 1‑29914‑5

Unknown

99‑4‑12

Erythema infectiosum

Polio‑ 1

Vaccine‑ like

99 ‑Fukushima‑ 1IY
2Y5Y

Unknown

98‑10‑30

99‑2‑ 1 7

99‑2‑1 7

書URD

Polio̲ 1Vaccine̲) ike

99 ‑ Fukushima127MM

None99141 I 5

99‑4‑30

99‑4‑3088ITP

Polio‑2

Vaccine‑like99 ‑Niigata‑ 33YF

9 7‑5 ‑9(ュnd)

99‑5‑1499‑5‑3 1

99‑5‑3 1URD

Polio̲ 1ⅥlCC ine‑like

9 9 ‑Ni igata‑47YFUnknown

99‑4‑ 1 3

99 ‑6‑24

99‑6‑25Herpanglna

Polio̲ 1

Vaccine‑ like

99‑Nara‑ 18MFNone

99‑1 0‑26

99‑1 1‑24

99‑1 1‑24Dia汀hea

Polio̲2& 3

Vaccine̲like

99 ‑Ni igata‑2* : Upper respiratory disease'': Idiopathicthrombocytopenic purpura*Corresponding author: Fax: +8 1 ‑42‑56 1 ‑4729, E‑mail:tyoneyam@nih.gojp90