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Pundyak Îleg ².
Institute of Ecology of the Carpathians of NAS of Ukraine
THE INFLUENCE OF GRAVITY ON THE POLARITY OF MOSS SPORES GERMINATION
Although today it have been given much attention to the investigations of the influence of gravity on primary growth directions of rhizoids of spores of fern Ceratopteris richardii (Chaterjee et al., 2000; Bushart, 2007), the peculiarities of gravidependent polarization of moss spores remain insufficiently studied. E. Heitz (1940; 1942) and H. Fitting (1950) established that in darkness gravity oriented germinating filaments of Funaria hygrometrica. Rhizoids grew downward and chloronema grew upward. L. Kofler and A. Claudet (1961) characterizing distributions of primary growth direction of the filaments in gravity field, made circular diagrams and also proposed mathematical criterion of geotropic response of the spores c:
c= ; (1.1)
Where ³ is a number of one of the sixteen sectors sharing the circle; is the percentage of the spores with the filaments growing within direction of sector i; is spatial orientation of sector i; N is quantity of investigated filaments. It was shown that the orientation of substrate surface strongly influence on the quantity ñ. Knowing nothing about these experiments, O. Pundyak with coauthors (2002 ) showed the influence of gravity on the polarity of F. hygrometrica spores germination. The circular diagrams of the filaments growth directions were made by means of the program especially created in objective oriented medium Delphi 5 using data bases Paradox 7, SQL form 2 requests. For the indication of gravisensitivity, the authors used the quantity of dispersion of the angles. The dispersion increased and the level of visual plastids sedimentation decreased, while the angle between substrate surface and the horizontal plane decreased. At the first time it was established that reorientation on 900 in vertical plane of the spores which have just formed rhizoids led to the formation of protonemal filaments growing at 900 to the rhizoids. The same phenomenon could be observed when the dishes with the spores were oriented horizontally in stable position.
The geotropic response was revealed also in several moss species. After F. hygrometrica the maximal ability to graviresponse were found in C.purpureus (Nurit, 1968). Therefore profound investigations and comparing the influence of gravity on polarity of F. hygrometrica and C. purpureus spores germination are logical prolongation of the field discovered by L. Kofler.
In 2004 Î. Pundjak proposed to quantify the spores gravisensitivity by criteria :
where is the percentage of the filaments growing within the interval of directions from i0 to j0; =0,28(j - i) is the percentage of the filaments, which could grow within this interval under the condition of the absence of the gravity influence.
Soon it was proposed criterion L: L = ln(a/b), where a and b are probabilities of the filaments growth upwards and downwards correspondingly. We established that L is proportional to the sinus of the dishes inclination angle. It approves our presumption that in F. hygrometrica and C.purpureus spores the thermal shiftings of plastids influence on polarization of the spores (Pund'ak, 2007 ). Thanks to natural spore heterogeneity, we showed experimentally that L linearly depends on dimension and quantity of plastids in a spore. We suppose that the spores polarize in consequence of the stretching of cytoskeleton structures, which support cell bodies. According to the investigation we can think that a spore polarize in the opposite direction (with rhizoids growing upward and chloronema filaments growing downward) in the case of simultaneous nanometric thermal dap of all the plastids in a spore, when the mechanical load on the cytoskeleton temporary disappear. As is known, thermal fluctuation may influence on system symmetry when it is found in the point of phase transition. The process of swelling of the spores after their sowing is connected with increasing of water content and therefore with increasing of thermal capacity of the spores in consequence of increasing of intramolecular motility of biopolymers. We assume that 24 hours after the sowing the spores were found in the point of phase transition of the second order. Thus, thermal fluctuations influence on the gravi-dependent polarization of the spores of moss F. hygrometrica (Pundiak, 2009 ).
The further decreasing of the inclination angle from 100 to 10 did not cause any changes of F. hygrometrica spores polarization in vertical direction, but the percentage of outgrowth threads growing horizontally considerably increased, as it was in diapason of the dishes inclination from 900 to 100. It can serve as confirmation of our hypothesis that the gravidependent polarization of the spores takes place from the beginning in horizontal direction and later in vertical thanks to plastids sedimentation (Pundyak, 2010).
L. Kofler (1971) showed, that in swelled spores of F. hygrometrica plastids sedimented, but vacuoles moved upward. Î. Pundjak and Î. Demkiv (2008 ) not only observed these phenomena, but made an attempt to give a quantitative and correlative characteristic of the plastids sedimentation, vacuoles flotation and the alternation of different growth directions of rhizoids and chloronema. It was shown, that rhythmical shiftings of the plastids and vacuoles correlate with callose distribution shiftings, which in one's turn correlates with the growth directions. According to L. Kofler (1971), the maximal response on geotropic stimulus on the stage of rhizoids formation was observed at 320Ñ. Our investigations complete the mentioned ones. Thus, we established for the first time that chloronemal filaments at these conditions grew not upward, but downward.
The spores' transfering from clinostat to stable position revealed decreasing but not blocking of rhizoids response on geotropic stimulus (Kofler, 1971). Our investigations complete the latter by the data concerning the spores' polarization on the stage of chloronema formation. As it was shown, after 24 hours cultivating of the spores on clinostat, immovable spores formed chloronemal filaments growing independently from the gravity. In 1962 L. Kofler with coauthors established, that glucose amplified the effect of geotropic response, but white light inhibited it. We investigated the influence of blue and red light on gravidependent polarization of F. hygrometrica and C. purpureus spores. It was revealed that red light blocks gravidependent polarization of both species, but blue light does it only for F. hygrometrica. Obviously in F. hygrometrica spores in geotropic response there involved pigment systems of phytochrome and cryptochrome, but in C. purpureus spores there involved only the phytochrome system. We also demonstrated, that before the germination of F. hygrometrica spores under the influence of polarized light, there appears the axis of gravisensitivity, along which a spore is able to perceive gravity. When such an axis was horizontally oriented by means of a polaroid, a spore was not gravisensitive (Pundjak, 2003). We also compared the influence of glucose on gravisensitivities of F. hygrometrica and C. purpureus spores. It was revealed that in contrast to F. hygrometrica, glucose inhibited gravisensitivity of C. purpureus spores (Pundyak et al., 2007). L. Cofler with coauthors (1962) demonstrated considerable influence of simultaneous action of vitamins thiamin, biotin, pyridoxin and nicotinic acid on geotropic response of F. hygrometrica spores. We showed the influence of EGTA (Pundiak, Demkiv, 2002), uridine diphosphate, calcium gluconate (Pundjak, 2004), lanthanum chloride (Pund'ak, 2006), nitrogenous oxide (NO) (Pundyak, 2009 ) and strong cooling (Pundyak, 2009 ) on such an characteristic. Thus, EGTA in concentration of two orders lower than that of calcium ions in nutrient medium inhibited the initiation of the primary rhizoids. As it is known, EGTA is concurrent inhibitor of glutamate receptors in plasma membranes of several organisms (Chen, 2000). F. hygrometrica spores evidently belong to this group. UDP, which is a product of glycosylation and activator of calcium channels, caused considerable decreasing of the influence of gravity on F. hygrometrica spores polarization. But in condition of the absence of calcium ions this effect was more essential. Lanthanum chloride, which is concurrent inhibitor of calcium channels in concentration of 60 mêM caused considerable increasing of influence of gravity on F. hygrometrica and C. purpureus spores polarization. We suppose that lanthanum ions influence on the spores polarization via cytoskeleton. We established that on the medium with glucose the influence of NO on F. hygrometrica spores polarization was mediated by guanylate cyclase activation: the action of nitroglycerine was the same with that of sildenafil and appears to increase the gravisensitivity. On the medium without glucose, NO influences on the gravisensitivity as antioxidant. It appears to decrease the gravisensitivity.
After strong cooling (at -260Ñ for 4 hours) F. hygrometrica spores forgot geotropic stimulus, but remembered the gravistimulus along horizontal direction. It can serve as the confirmation that the polarization takes place in the state of phase transition of the second order (Pundiak, 2009) and this process can be modeled by the movement of virtual particle along vertical meridian of a swelled spore (Pundiak, 2008).
1. Kofler L. & Claudet A. Sensibilite geotropique des germinations de Funaria hygrometrica (L.) Sibth.// Rev. Bryol. et Lichen.- 1961. - 30. - P. 13-20.
2. Kofler L., Dutel J. & Nurit F. Variations in the geotropic sensitivity of germinating Funaria spores in response to some external influences// J. Linn. Soc. (Bot.) - 1962. - 58. - P. 311-319.
3. Kofler L. (1971) Influence de la temperature sur le geotropisme des spores de Funaire // C.R.Acad.Sc.Paris. - 1971. - t. 272. - P.1244 - 1247.
4. Nurit F. Germination et sensibilite geotropique des spores de quelques especes de Mousses// Extrait de la revue Briologique et Lichenologique. -1968. - t. 36, f. 1-2. - P. 148-154.
5. Pundyak O. I., Demkiv O. T., Khorkavtsiv O. Ya, Bagrii B. B. Polarity of Spore Germination in Funaria hygrometrica Hedw.// Kosmichna Nauka i Tekhnologiya. - 2002. - t. 8, No. 1. - P. 96 - 100. http://adsabs.harvard.edu/abs/2002KosNT...8a..96P
6. Pundiak O., Demkiv Î. Temporaly germinating rhythms of moss Funaria hygrometrica Hedw. spores. //COSPAR. 02-A-01942 (2002).
7. Pundjak O. Investigation of the Influence of Polarized Light on the Gravisensitivity of Funaria hygrometrica Hedw. Spores// Journal „Scientific principles of conservation of biotic diversity" - 2003. - Issue 5. - P. 298 - 302. http://www.ecoinst.lviv.ua/b5-2003/fbs5.pdf
8. Pundiak O.I. The Influence of Modulators of Calcium Exchange on the Gravisensitivity of Spores of Moss Funaria hygrometrica Hedw.// Journal „Scientific principles of conservation of biotic diversity".- 2004. - Issue 3, - P. 156-161. http://www.ecoinst.lviv.ua/b6-2004/fbs1.pdf
9. Pund'ak O.I. Investigation of Moss Funaria hygrometrica Hedw. Spores Germination in the Gravity Field// Journal „Scientific principles of conservation of biotic diversity".- 2006. - Issue 7. - P. 190-195. http://www.ecoinst.lviv.ua/b7-2006/fbs8.pdf
10. Pund'ak Î.². Gravi-dependent polarization spores of moss Funaria hygrometrica Hedw.// Uzhhorod University Scientific Herald. Series Physics. - 2007. - Issue 21. - P. 186-190. // http://www.univ.uzhgorod.ua/tte/images/visnyk/21/34.pdf
11. Pundyak O., Demkiv O., Khorkavtsiv Ya. Comparing the gravisensitivities of spores of mosses Funaria hygrometrica and Ceratodon purpureus //Matherials of Conferention «The Problems of studying and protection of biodiversity of Carpathians and its environs», ²vano-Frankivs'k, 2007. - P. 96-98. http://www.nbuv.gov.ua/portal/natural/vpnu/2007/07-08.pdf
12. Pundiak O. A model of gravisensitivity of moss spores // Materials of Conference «Contemporary Problems of applied Mathematics».- 2008. - L'viv.- P. 36. http://blues.lnu.edu.ua/ami/billtexts/tezy_SPPMI2008.pdf
13. Pundjak O., Demkiv O. Investigation of Gravity Dependent Plastides and Vacuoles Distribution in Germinating Spores of Moss Funaria hygrometrica Hedw.// Scientific Society of T. Shevchenko Gerald. - 2008. - Issue 23. - P. 156 - 162. http://www.nbuv.gov.ua/portal/natural/pntsh/ekolzb/2008_23/PUNDJAK.pdf
14. Pundiak O.I. Gravi-dependent polarization of germinating spores of moss Funaria hygrometrica Hedw // Materials of Conference "²ÅF-2009" (Uzhorod, 2009) - P. 56.
15. Pundyak Î.². Investigation of NO Influence on Gravisensitivity of spores of moss ìîõó Funaria hygrometrica Hedw.// Matherials of Ukrainian Internet Conference. - Kyjiv, 2009. - P. 4-6. http://intkonf.org/pundyak%20-oi-doslidzhennya-vplivu-oksidu-azotu-no-na-gravichutlivist-spor-mohu-funaria-hygrometrica-hedw/
16. Pundyak Î.². The influence of cooling of gravistimulated moss spores on the polarity of their germination// Matherials of Ukrainian Internet Conference. - Kyjiv, 2009. - P. 14-16. http://intkonf.org/pundyak-oi-vpliv-oholodzhennya-gravistimulovanih-spor-mohu-funaria-hygrometrica-hedw-na-polyarnist-yih-prorostannya/
17. Pundyak Î.². Investigation of the influence of the substrate surface orientation on polarization of spores of moss Funaria hygrometrica Hedw. // Matherials of Ukrainian Internet Conference. - Kyjiv, 2010. http://intkonf.org/pundyak-oi-doslidzhennya-polyarizatsiyi-spor-mohu-funaria-hygrometrica-hedw-zalezhno-vid-orientatsiyi-poverhni-substratu/