Chemistry: natural resins or derivatives; peptides or proteins; – Proteins – i.e. – more than 100 amino acid residues
Reexamination Certificate
2002-01-23
2004-11-30
Wax, Robert A. (Department: 1653)
Chemistry: natural resins or derivatives; peptides or proteins;
Proteins, i.e., more than 100 amino acid residues
Reexamination Certificate
active
06825321
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to plant thermogenic genes and proteins. More particularly, the invention relates to thermogenic genes derived from a skunk cabbage (
Symplocarpus foetidus
) and gene products (proteins). Those genes and proteins are useful in breeding of cold-avoidance plants, medical treatment of diabetes metllitus or obesity, or development of novel thermogenic bio-materials.
2. Description of the Related Art
Stresses due to low temperatures, droughts and salinity are common harmful environmental factors that terrestrial plants encounter. Among these stresses, it has been considered that cellular injury due to the low temperature is the most important factor which restricts productivity of crops (Levitt, 1980). To resist the low temperature stress, cold-hardy plants such as wheat or rye have a variety of physiological and metabolic responses which lead to cold acclimation (Sakai and Larcher, 1987; Steponkus, 1984; Thomashow, 1998; Uemura and Steponkus, 1997). In contrast, it is known that some plants including skunk cabbage have a specialized system by which the plants generate heat to avoid freezing (Knutson, 1974; Nagy et al., 1972; Schneider and Buchenen, 1980).
The temperature of the flower in the spadix of skunk cabbage, which flowers in early spring, has been known to maintain its temperature at higher than +10° C. even when the ambient temperature falls to −15° C. (Knutson, 1974). For example, thermoscopic analysis using infrared camera indicates homeothermic behavior of the surface temperature of the spadix (FIG.
1
). It should be noted that, in this experiment, the plants were placed in the growth chamber and the air temperature was gradually decreased. As clearly seen from
FIG. 1
, the temperature of the spadix of skunk cabbage is kept at approximately 19° C. notwithstanding a fall of the ambient temperature.
The temperature is thus maintained by doubling the respiration rate from the level of 12° C. to that of sub-zero temperature. It has also been considered that the heat production in thermogenic plant species relates to a cellular metabolism called cyanide-non-sensitive
on-phosphorylating electron-transferring pathway, which is controlled by mitochondrial alternative oxidase (AOX)(Berthold and Siedow, 1993, Ito et al., 1997; McIntosh, 1994, Wangner and Krab, 1995).
On the other hand, it has been shown that a mitochondrial protein called an uncoupling protein (UCP) plays an important role in generation of heat in mammals. UCP found in the intima of mitochondria make H
+
flow into the membrane to uncouple aspiration from synthesis of ATP which acts to disperse chemical energy to metabolic heat (Klaus et al., 1991; Kihngeberg and Winkler, 1985; Ricquier et al., 1991). In animals, 3 types of UCPs have been found. UCP1 is primarily distributed in brown adipose tissue (Nichollus and Locke, 1984). UCP2 is found ubiquitously in many tissues (Fleury et al., 1997), and UCP3 is localized specifically in skeletal muscle (Boss et al., 1997).
It has been considered that UCPs of mammals, similarly to other carrier proteins of mitochondria, are composed of 6 transmembrane segments, of which the hydrophobic portion is derived from pairing amphipathic &agr;-helix structure (Liu et al., 1988; Maia et al., 1998). It is also known that the activity of these UCPs decreases depending on purine nucleotides (ATP, GTP, GDP and ADP) attached to the C-terminal legion and increases by free fatty acids (Jezek et al., 1998; Lin and Klingenberg, 1982; Katiyar and Shrago, 1989; Rial et al., 1983; Sluse et al., 1998).
On the contrary, 2 cDNAs encoding UCP-like proteins of plant origin were isolated from potato (StUCP: Laloi et al., 1997) and from Arabidopsis (AtPUMP: Maia et al., 1998). Since the expression of StUCP was mainly detected in the flower and the Suit, it has been postulated that StUCP may concern respiration during flowering and maturation of the fruit together with the AOX activity (Laloi et al., 1997).
Potato and Arabidopsis have been considered to be non-thermogenic plants. However, the expression of StUCP and AtPUMP have been induced by low temperature. Therefore, it has been suggested that these genes are involved in heat production (Laloi et al., 1997; Maia et al., 1998).
In the thermogenic plants such as skunk cabbage, however, UCP-mediated thermoigenic mechanisms have not yet been identified.
The purpose of the invention of this application is to provide unidentified novel UCP genes derived from a thermogenic plant, skunk cabbage.
The additional purpose of this application is to provide skunk cabbage UCPs which are expression products of the novel genes.
SUMMARY OF THE INVENTION
The invention provides thermogenic genes derived from skunk cabbage, i.e., gene SfUCPa of which cDNA comprises the base sequence of SEQ ID NO: 1, and gene SfUCPb of which cDNA comprises the base sequence of SEQ ID NO: 3.
Moreover, the invention provides thermogenic proteins, i.e., protein SfUCPA expressed from SfUCPa, which comprises the amino acid sequence of SEQ ID NO: 2, and protein SfUCPB expressed from SfUCPb, which comprises the amino acid sequence of SEQ ID NO: 4.
In addition, the invention provides cDNA having the base sequence of SEQ ID NO: 1 or a partial sequence thereof, and cDNA having the base sequence of SEQ ID NO: 3 or a partial sequence thereof.
REFERENCES:
patent: 6602694 (2003-08-01), Albrandt et al.
patent: WO 9632483 (1996-10-01), None
Ito, K., “Isolation of two distinct cold-inducible cDNAs encoding plant uncoupling proteins from the spadix of skunk cabbage (Symplocarpus foetidus)”, Plant Science, Dec. 1999, vol. 149, No. 2, pp. 167-173.
Ito, K. “A cold-inducible gene encoding uncoupling protein in thermogenic plant species” Cryobiology and Cryotechnology, Dec. 1999, vol. 45, No. 2, pp. 43-46.
Ricquier, D. et al., “The uncoupling protein homologues: UCP1, UCP2, UCP3, StUCP and AtUCP”, Biochem. J., 2000, vol. 345, No. 2, pp. 161-179.
Watanabe, A. et al., “AtUCP2: a novel isoform of the mitochondrial uncoupling protein of Arabidopsis thaliana” Plant Cell Physiol., Nov. 1999, vol. 40, No. 11, pp. 1160-1166.
Maia I.G. et al., “AtPUMP: an Arabidopsis gene encoding a plant uncoupling mitochondrial protein”, FEBS Letter, 1998, vol. 429, pp. 403-406.
Laloi, M. et al., “A plant cold-induced uncoupling protein”, Nature, 1997, vol. 389, pp. 135-136.
Japan Science and Technology Corporation
Wenderoth , Lind & Ponack, L.L.P.
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