RNA with a high cytosine and a low guanine content in poliovirus infected cells

Vol. 15, No. 4, 1964

BIOCHEMICAL

RNA WITH

AND BIOPHYSICAL RESEARCH COMMUNICATIONS

A HIGH CYTOSINE IN POLIOVIRUS G.

Institut

Koch

AND A LOW GUANINE! INFECTED CELLS

and

H.

CONTENT

Kubinski

zur

Erforschung der spinalen und der Multiplen Sklerose Universitltskrankenhaus Hamburg-Eppendorf,

Kinderlahmung Germany

Received February 24,1%4 Infection crease

in

1962). (to

of the

rate

be

after

RNA

infection of

is

albumin

column

analysis

of

RNAs. virus

of

with

a high

This

RNA

RNA

infected

animal

Georgiev

et

concentrations (Cr-RNA)

is

by

et

can

1962)

of

labeled

(C)

the

and

the virus

(G)

onset

content. an

messenger

C-rich, infected

308

the

RNA in

Darnell,

ribosomal

cel-

weight

to

presence

actinomycin,

us

composition

as and

by

subsequent

before

guanine

act

(Scherrer

in

32 P-labeled

viral

comparable

In

of

RNA enabled

a nucleotide

to

ribosomal

a molecular

low

the

of

and

recognized

and

1963).

contamination

poliovirus

from

has

two

a methyl-esterified

al.,

be

supposed

al.,

within

especially

on

a de-

inhibition

that

cells

cells

RNA and

cells

this

Fractionation

it

synthesized no

of

shown

properties

of is

or

viral

possesses which

(Holland,

RNA distinguishable

cytosine

fraction

synthesis

RNA and

RNA production; to

RNA

composition

This

comparable

causes

human

(Kubinski

an

poliovirus

have

infected

base

by

mechanism

impaired.

from

identify

the

messenger

isolated

lular

total

elsewhere)

RNA precursor

little

of on

reported

synthesis

of

cells

Experiments

hours

to

animal

non-

1962; of

moderate G-poor

cells precursor

RNA with RNA.

Vol. 15, No. 4, 1964

BIOCHEMICAL

Amnion were

cells

(or

in

washed

phosphate-free

actinomycin

(2

50 PFU of

were

by

RNA was on

composition

termined.

The

used

et

1962;

Figure

la

an

the cell

two

reported

The high

was

Fraction RNA.

never

II The

base

fraction

I

in

composition similar

fraction

II

distinctly

differs

from

messenger"

RNA

In

presence

the

in

the

Cr-RNA

is

(Table

inhibition

(figure

lb)

and

and

to has

that

of

RNA

pre-

RNA fractions,

as

1963 b).

Frac-

virus

RNA.

infective

of

hours

enters

infective

1)

obtained

3.5

for

poliovirus

32 P-labeled

of

ribosomal

a base

composition

polio

RNA and

both

in With

of no

(Table

de-

1964).

Koch,

to

in

de-

RNA

RNA.

The

which "cellular

1).

formed.

overall

size

The

was

label

contain

and

column.

incubated

and

is

32 P-RNA

diagram

(Kubinski

corresponds

phenol

al.,

radioactive

to

samples

described

et

weight

found

hot

been

molecular

previously I

elution

was

RNA fractions

Koch

culture,

actinomycin.

dominantly

tion

shows

infected

without

al.,

have

with

times

albumin the

cells/

without

medium

various

with

of

methods

(Xubinski

from

extracted

and

the

At

x 106

infected

later

a methyl-esterified

32 P-nucleotide

tail

minutes

with

were

radiophosphate.

withdrawn,

fractionated

cells

suspension

(4

medium

The 40

in

resuspended

Eagle's

and

RESEARCH COMMUNICATIONS

incubated

and

ug/ml).

virus

supplemented

in

HeLa-cells)

centrifuged,

ml)

AND BIOPHYSICAL

the

absence actinomycin,

RNA labeling

synthesis

of

of

tected.

309

actinomycin however,

was ribosomal

as

high

D, the

as

RNA was

97% de-

Vol. 15, No. 4, 1964

BIOCHEMICAL

AND BIOPHYSICAL RESEARCH COMMUNICATIONS

Fraction

FIGURE Chromatographic cells

separation

infected

for

3.5 la) lb)

hours

at

without 2 ug 12

-o-o-c

with

50

period.

After

sized.

Cr-RNA

is

from

poliovirus

amnion

and

incubated

8 UC of D added

at

32P/ml time

of

infection;

32P/ml

W absorbance: 32 P (please

Cr-RNA

of

obtained

36'~.

actinomycin

UC of

RNA

PFU/ml

actinomycin;

and

The

of

1

A 260 note

that

the

scale

differs

in

fig.

b)

synthesized the starts

eclipse to

exclusively only elute

poliovirus from

310

within

the

RNA column

the

eclipse

is

synthe-

at

lower

la

BIOCHEMICAL

Vol. 15, No. 4, 1964

AND BIOPHYSICAL RESEARCH COMMUNICATIONS

TABLE $ Distribution

1

32 P-nucleotides

of

in C

* ribosomal fraction

I,

fraction

II,

la

figure figure

polio* "cell

*

mean

salt

values

obtained

and

to

the

unstable

The

role

fected

of

cells

is

formation

is

has

an

the

base

same

the

that

and

it

less

mycin

in

be host is

1963 reduced

than

that

26.9

20.0

21.2

22.5

28.3

25.7

23.6

21.8

26.1

28.0

23.2

34.0

28.1

20.1

17.8

experiments.

It the

virus

of

fact

since

they

the

shown

(Georgiev

that

the

by

moderate

of

ribosomal

virus

from

that

actinomycin

both

to

RNAsare

the

column

at

explana-

of

is

Cr-RNA of

cellular

di-

Cr-RNA

sug-

"messenger" al.,

synthesis

1963; of

concentrations RNA precursor. 311

its

plausible

properties

et

in-

Moreover,

that

production

the

in

complementary

A more

to

figure

b).

unlikely

not

elute

The

1963 of

in

earlier

synthesis.

spite

cell.

I

RNA since

its

related

a)

Koch,

seems

is

the

fraction

observed

Cr-RNA

that

recently

Koch,

32.0

metabolism

concentration.

by

32.9 28.5

and

on

of

size,

to

was

by

salt

20.0 22.2

to

the

effect

seems

rected gest

directed

RNA

same

tion

known.

composition

the

the

in

not

inhibitory

poliovirus of

Cr-RNA

G

17.2 19.4

several

(Kubinski

U

29.9 29.8

RNA fraction

cells

RNA fractions

A

corresponding

polio-infected

is

in

concentrations,

la

It

la

*

messenger"

*

,,crtt

various

RNA.

Kubinski

messenger of

RNA actino-

Vol. 15, No. 4, 1964 Two

possibilities

thesis

of

lular so

BIOCHEMICAL

some

RNA that

is

G-rich,

newly

C than

in

sis

a special

second

might

G;

or

(2)

viz.

that

virus

messenger

can

the

of

One induces

labeled the

This that

a new

is

the

in

synthe-

some

protein,

1957).

could

cel-

infection

finding

interferon

1963).

virus

period.

Lindenmann, of

syn-

weight

induce

eclipse by

the

strongly

synthesis

and

infection

Cr-RNA

by

more

the

the

It

is

inhibited

expect,

by

therefore,

formation

of

a new

interferon.

investigations

of

molecular

infection

production

RNA for

Further

is

supported

(Heller,

the

role

the

(Isaacs

actinomycin that

RNA

trigger

the

(1)

inhibited

is

interferon

known

high

RNA during

infections

considered:

C-poor

formed

possibility

virus

be

preferentially

the

of

AND BIOPHYSICAL RESEARCH COMMUNICATIONS

in

are

called

poliovirus

for

infected

to

clarify

the

cells.

Acknowledgement The

expert

technical

gratefully

assistance

acknowledged.

ported

by

schung

and

of

This

Bundesministerium Deutsche

Miss

M.

investigation

fiir

Sichart was

is sup-

wissenschaftliche

For-

Forschungsgemeinschaft.

References Georgiev, and

G.P., Severtzov,

Heller, Holland,

E.

Koch,

G., (1964).

Virology

J.J.

Isaacs, A. (1957).

Samarina, A.N. 21:

Drees,

Lindenmann, 0.

65:~

Lerman, M.I., Smirnov, ZOO: 1291 (1963).

and

M.N.

(1963).

Biophys.Res.Commun.

Biochem. and

O.P., Nature

J.

9:

Proc.Roy.Soc.

Kubinski,

312

H.

Z.

556

(1962).

B 147: Naturf.,

258 in

press

Vol. 15, No. 4, 1964

BIOCHEMICAL

AND BIOPHYSICAL

RESEARCH COMMUNICATIONS Tagung

Kubinski, 19.

H. and - 21.9.1963

Koch, a).

G.

Biochemische

Kubinski,

H.

Koch,

G.

J.Mol.Biol.

and

Drees.

Kubinski, 61:

332

Mandell, Scherrer, 7: 486

and

H., Koch, (1962).

J.D.

and

K. and (1962).

G. Hershey,

Darnell,

A.D. J.E.

6: 0.

102

Strasbourg, (1963)

Biochim.Biophys.Acta

Anal.Biochem.

1:

Biochem.Biophys.Res.Commun.

313

b).

66

(1960).