Bud formation inHydra: Inhibition by an endogenous morphogen

Abstract

From crude extracts ofHydra tissue a substance has been purified which prevents or retards the asexual reproduction by budding. The molecular weight is in the range of 300 to 1000 daltons. Inhibition of bud formation can be observed with concentrations equivalent to the extract from one hydra per 4 ml, that is, to a more than 10,000-fold dilution of the initial crude extract of a hydra. The purified inhibitor is active at a concentration of less than 10 M.Most of the inhibitor present inHydra is bound to cells. Within the cells the substance is mainly bound to particulate structures which sediment at 10,000 g. Its concentration is highest in the hypostomal region and decreases in the direction of the tentacles and peduncle. A second, lower, peak has been found in the basal disc. Treatment of the animals with a toxic agent (nitrogen mustard) which depletes the animal of interstitial cells, nematocytes and nematoblasts excludes the possibility that the inhibitor is present to any great extent in these cells. In conjunction with cell separation experiments by centrifugation of fixed cells in suspension, these results indicate that nerve cells are the most likely sites of storage of the inhibiting substance, although epithelial cells are not excluded as sources for the inhibitor.

Pattern of epithelial cell cycling in hydra

“…Berking (1974Berking ( ,1977Herrmann and Berking, 1987) has described a factor, inhibitor I, which blocks mitosis of epithelial cells. Treatment of hydra with the factor inhibits mitosis almost immediately, indicating that the factor acts late in G2 phase.…”

Methionine in pattern control of Hydra

“…This fraction was passed through anion and cation exchangers, at neutral pH [BioRex 70 (H + -form); Dowex AG1 (OH + -form), BioRad]. Up to this point the purification was almost identical to that described earlier [12,13,14]. The experiments done and published at that time can therefore be related to the data presented here.…”

“…Head regeneration was studied in budless animals one day after the last feeding [2,[12][13][14][15]. The animals were sectioned at about one half of the body length and the aboral (foot-bearing) fragments were studied.…”

“…In head-regenerating animals new tentacles became visible the following day. The ability of foot-regenerating animals to attach to a surface was used to test for the presence of a new foot [2,[12][13][14][15]. Animals regenerating a foot were transferred to new dishes with fresh culture medium (after one washing) 22 hours after sectioning and sampling from the dishes.…”

“…One active fraction was detected [12,13,14]. Following application of various separation methods (see Experimental Procedures) one still-active fraction was obtained [14].…”