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“Membrane knotting” between blastomeres of limnea
Isolated cilia from Tetrahymena pyriformis
631
and analysed. The preparations, 30-40 mg dry wt, gave a strong positive colour test 131 for protein. The composition of an hydrolysate (5.8 N HCI, 108”C, 21 hours) was determined by ion-exchange chromatography [4]; the results are given in Table I. An unknown ninhydrin-positive substance emerged from the ion-exchange columns in between cysteic acid and methionine sulphoxide. The amino acid residues accounted for about 70 per cent of the mass of the dry, ash-free preparation. A fraction of the preparations (15-25 per cent by wt) was soluble in chloroform-methanol (2:l). The anthrone colour reaction for polysaccharides in the cilia was equal to only 1 per cent of that given by an equal weight of glucose, and ultraviolet absorption measurements 121 showed that only 0.3 per cent of ribonucleic acid was present. \Ve wish to thank
Dr. H. B. Fell, F.R.S.
for counsel during
these experiments.
REFERENCES
1. CCI~LD, F. JI., Ezptl.
Ce// Research 18, 258 (1959). W. A. and ROSSITER, R. .J., Hiochem. .J. 51, 480 (1952). 3. LOWHY, 0. H., Rosmnouc~, N. .J., FARR, A. L. and RANDALL. R. J., .J. Viol. 265 (1951). 4. MOORE, S., SP.ACIMAN, I>. H. and STEIN, \V. H.. AIIU/. Chem. 30, 118.5 (1958).
2. LOGAS, J. E.,
MASNELL,
“MEMBRANE
KNOTTING”
BETWEEN
Chrm.
193,
BLASTOMERES
OF LIMNEA C. H. WADDINGTON, Institute
of Animal
Genetics,
M, M. PERRY University
and E. OKADA
of Edinburgh,
Scotland
Received February 15, 1961
IN discussing the adhesion of blastomeres in early sea-urchin embryos, Balinsky [lj has listed three mechanisms by which cells appear to stick together. (i) In “ordinary contact”, long stretches of two plasma membranes remain in relatively close contact, but with a gap of some 140 A between them, this being presumably filled by some electron-light intercellular substance. (ii) In some cases contiguous cells seem to be held together by the formation of numerous interdigitating processes. (iii) Intercellular connecting bars appear, usually in groups forming the structures known as desmosomes. In the cleavage stages of the eggs of the mollusc Limnea peregra we have seen what appears to be a new form of inter-cellular “contact body”. During early cleavage the blastomeres are not everywhere in close contact with one another in the centre of the embryo, although a well-developed blastocoel has not yet been formed. In the angles of the spaces between cells, or occasionally bridging across the middle of a gap, one sometimes finds ovoid or roughly spherical bodies, which are made up of highly Experimental
Cell Research
23
C. H. Waddington, M. M. Perry and E. Okada
Figs. l-4.-“Membrane Knots” (M.K.) between blastomeres in Limnea peregra embryos. N, granules”, (lipid ?). nucleus; 8, j3-granules of yolk containing crystalline protein; W, “wheel-shaped All x 7,800. Experimental
Cell Research 23
‘cAfembrane knotting”
633
between blastomeres of Limnea
folded regions of the cell membranes. The overall dimensions of the bodies measure one or two mp, and a straight line through one of them would cross some 8-15 membrane profiles. In some cases it is difficult to be sure that the membranes which are folded together are derived from more than one cell, but in several examples this seems fairly certain, and it is thought that in general the bodies represent places in which the membranes of two (or more) cells are crumpled together to form what may be could play an important part in regarded as a “knot”. Presumably such “knots” holding the cells together. In these eggs we have not seen any desmosomes, but they have not been searched for at high magnification. Elbers ([3], Pl. 7) has illustrated what appear to be desmosomes in the eggs of Limnen stagnatis at a similar stage, but does not appear to have seen any membrane knots in this species. It would, perhaps, be unsafe to attach too much weight to this negative evidence: possibly demosomes exist in L. peregra and membrane knots in L. stagnatis, but have in both cases so far escaped detection. On the other hand, there is another indication that the eggs of the two species may differ rather radically. In both one finds mitochondria and yolk granules containing crystalline aggregates of protein (p granules [2]); but in L. peregra there are in addition many very characteristic “wheel-shaped granules”. These are very numerous in certain regions of the egg, in the early cleavage stages. They do not seem to occur in this form in L. stagnatis; possibly they correspond to the “lipid droplets” which fix as homogeneous spheres in that form ([3], Pl. 15, 16). REFERENCES 1. RALISSI~Y, U. I., Exptl. Cell Research 16, 429 (1959). 2. ELBERS, P. b’., Koninkl. Nederl. Akad. Tl’etenschap C. 60, 96 (1957). 3. ELBERS, P. l:., Over de beginoorzaak van het L-effect in de Jlorphogenese. Utrccht, 1959.
Experimental
Ph.D.
thesis,
Cell Research 23
631
and analysed. The preparations, 30-40 mg dry wt, gave a strong positive colour test 131 for protein. The composition of an hydrolysate (5.8 N HCI, 108”C, 21 hours) was determined by ion-exchange chromatography [4]; the results are given in Table I. An unknown ninhydrin-positive substance emerged from the ion-exchange columns in between cysteic acid and methionine sulphoxide. The amino acid residues accounted for about 70 per cent of the mass of the dry, ash-free preparation. A fraction of the preparations (15-25 per cent by wt) was soluble in chloroform-methanol (2:l). The anthrone colour reaction for polysaccharides in the cilia was equal to only 1 per cent of that given by an equal weight of glucose, and ultraviolet absorption measurements 121 showed that only 0.3 per cent of ribonucleic acid was present. \Ve wish to thank
Dr. H. B. Fell, F.R.S.
for counsel during
these experiments.
REFERENCES
1. CCI~LD, F. JI., Ezptl.
Ce// Research 18, 258 (1959). W. A. and ROSSITER, R. .J., Hiochem. .J. 51, 480 (1952). 3. LOWHY, 0. H., Rosmnouc~, N. .J., FARR, A. L. and RANDALL. R. J., .J. Viol. 265 (1951). 4. MOORE, S., SP.ACIMAN, I>. H. and STEIN, \V. H.. AIIU/. Chem. 30, 118.5 (1958).
2. LOGAS, J. E.,
MASNELL,
“MEMBRANE
KNOTTING”
BETWEEN
Chrm.
193,
BLASTOMERES
OF LIMNEA C. H. WADDINGTON, Institute
of Animal
Genetics,
M, M. PERRY University
and E. OKADA
of Edinburgh,
Scotland
Received February 15, 1961
IN discussing the adhesion of blastomeres in early sea-urchin embryos, Balinsky [lj has listed three mechanisms by which cells appear to stick together. (i) In “ordinary contact”, long stretches of two plasma membranes remain in relatively close contact, but with a gap of some 140 A between them, this being presumably filled by some electron-light intercellular substance. (ii) In some cases contiguous cells seem to be held together by the formation of numerous interdigitating processes. (iii) Intercellular connecting bars appear, usually in groups forming the structures known as desmosomes. In the cleavage stages of the eggs of the mollusc Limnea peregra we have seen what appears to be a new form of inter-cellular “contact body”. During early cleavage the blastomeres are not everywhere in close contact with one another in the centre of the embryo, although a well-developed blastocoel has not yet been formed. In the angles of the spaces between cells, or occasionally bridging across the middle of a gap, one sometimes finds ovoid or roughly spherical bodies, which are made up of highly Experimental
Cell Research
23
C. H. Waddington, M. M. Perry and E. Okada
Figs. l-4.-“Membrane Knots” (M.K.) between blastomeres in Limnea peregra embryos. N, granules”, (lipid ?). nucleus; 8, j3-granules of yolk containing crystalline protein; W, “wheel-shaped All x 7,800. Experimental
Cell Research 23
‘cAfembrane knotting”
633
between blastomeres of Limnea
folded regions of the cell membranes. The overall dimensions of the bodies measure one or two mp, and a straight line through one of them would cross some 8-15 membrane profiles. In some cases it is difficult to be sure that the membranes which are folded together are derived from more than one cell, but in several examples this seems fairly certain, and it is thought that in general the bodies represent places in which the membranes of two (or more) cells are crumpled together to form what may be could play an important part in regarded as a “knot”. Presumably such “knots” holding the cells together. In these eggs we have not seen any desmosomes, but they have not been searched for at high magnification. Elbers ([3], Pl. 7) has illustrated what appear to be desmosomes in the eggs of Limnen stagnatis at a similar stage, but does not appear to have seen any membrane knots in this species. It would, perhaps, be unsafe to attach too much weight to this negative evidence: possibly demosomes exist in L. peregra and membrane knots in L. stagnatis, but have in both cases so far escaped detection. On the other hand, there is another indication that the eggs of the two species may differ rather radically. In both one finds mitochondria and yolk granules containing crystalline aggregates of protein (p granules [2]); but in L. peregra there are in addition many very characteristic “wheel-shaped granules”. These are very numerous in certain regions of the egg, in the early cleavage stages. They do not seem to occur in this form in L. stagnatis; possibly they correspond to the “lipid droplets” which fix as homogeneous spheres in that form ([3], Pl. 15, 16). REFERENCES 1. RALISSI~Y, U. I., Exptl. Cell Research 16, 429 (1959). 2. ELBERS, P. b’., Koninkl. Nederl. Akad. Tl’etenschap C. 60, 96 (1957). 3. ELBERS, P. l:., Over de beginoorzaak van het L-effect in de Jlorphogenese. Utrccht, 1959.
Experimental
Ph.D.
thesis,
Cell Research 23