The presence of two types of β‐cyanoalanine synthase in germinating seeds and their responses to ethylene

Abstract

The presence of two types of /^-cyanoalanine synfhase in germinating seeds and fheir responses to ethylene. -Physiol. Plant. 93: 71.3-718.Germinafine seeds of many species contain fwo fypes of /i-cyanoalanine synthase (CAS. EC 4.4.1.9) thaf conveii HCN fo /?-cyanoa'lanine. One is cyfoplasmic CAS (cyt-C.AS), which is precipitated by 50 lo 609r (NHai^SO^ and has a pH optimum of 10.5. Cvtoplasmic C.^S is present at high levels in dry seed and its activity does nof increase during imbibition. The activity ot cyt-CAS is uot affected by exogenously applied efhylene (CjHji. except in riee (Or\~a sativa cv. Sasanishiki). The second fype of CAS ftiuud in seed is mitochondrial CAS (mit-CAS I. which is precipifated by 60 fo 1()''« (NHjl:SO4 and has a pH optimum of 9.5. Mitochondrial CAS is present at low levels iu dry seed, and its activity increases greatly during imbibition in the seeds of ali species fested. Exposure fo C:H^ stimulated mit-C.i^S activity in seeds of rice, barley (Hordeum yidgare cy, Hadakamugii, cucumber (Cucumis satiyus cv. Kagafushinari) and cocklebur {Xanihhmi pennsyhamcum). The increase in fhe mit-CAS activity in cocklebur in response to C^Hj ccmimeticed after a fag period of 2 to 3 h when fhe duration of soaking was short (16 hi. but commenced without a lag period when fhe seeds were soaked lor Ihree mouths. Application ot bofh chioramphenicol aud cycloheximide to the axial and eotyledonary tissues of cocklebur seeds strongly inhibifed growth as well as fhe increase in mit-CAS acfivity. It is postulated fhal the mit-CAS is syuthesized de noi-o dtiring imbibition and thai ifs acfivity is regulated by C;H4. CO;, vvhich also promotes seed genninatiou in some species. v»as ineffective in stimulating tnif-C.AS activity iu cocklebur seeds.

Cytosolic r -Cyanoalanine Synthase Activity Attributed to Cysteine Synthases in Cocklebur Seeds. Purification and Characterization of Cytosolic Cysteine Synthases

“…Eighty percent of the total cytosolic CAS activity is precipitated by 40-70% saturation with (NH 4 ) 2 SO 4 ; the solubilized mitochondrial CAS activity is precipitated at 50-80% saturation (data not shown). The apparent optimum pH of each CAS activity differed, with the former at 10.5 and the latter at 9.5 (Hasegawa et al 1995b). Moreover, the two CAS activities were clearly separated by non-denaturing PAGE.…”

“…In contrast, the mitochondrial CAS activity was divided into two bands, one of which appeared on a gel at a position similar to that of the cytosolic one (compare lane 1 and 4). In a previous paper, we reported that the CAS activity of cocklebur seeds was increased by ethylene (Hasegawa et al 1995b). This promotion by ethylene was also indicated on the gels by the different strengths of the staining activity (compare lanes 1 and 4 with lanes 2 and 3).…”

“…However, CAS activity was also found in the soluble or cytosolic fraction from many plant tissues (Cooney et al 1980, Ikegami et al 1989, Manning 1988. Recently, we detected CAS activity in the cytosolic fraction as well as in the mitochondrial fraction in cocklebur seeds (Hasegawa et al 1995b). The optimum pH and the concentrations of ammonium sulfate required for precipitation differ between the CAS activity in the mitochondrial and cytosolic fraction.…”

A soluble β-cyanoalanine synthase from the gut of the variegated grasshopper Zonocerus variegatus (L.)

“…The enzyme from White birch, Betula verrucosa, was reported as a 38 Kd homodimer (Vahala et al, 2003). In summary, b-CAS has been found to exist in two molecular forms: a 52 Kd homomer and a homodimer with subunit sizes of 28-30 Kd (Hendrickson and Conn, 1969;Ikegami et al, 1988a, b;Hasegawa et al, 1995).…”

“…b-CAS has also been found to exist in two molecular forms: a 52 Kd homomer and a homodimer with subunit sizes of 28-30 Kd (Hendrickson and Conn, 1969;Ikegami et al, 1988a, b;Hasegawa et al, 1995). Another enzyme, cysteine synthase (O-acetylserine (thiol) lyase; CS or O-ASTL; EC 4.2.99.8), which exists in three isofroms: cytosolic (cysteine synthase A), chloroplastic (cysteine synthase B) and mitochondrial cysteine synthases, has been found to be involved in cyanide detoxification alongside b-CAS in higher plants such as in spinach and in Arabidopsis Hawkesford, 1998, 2000).…”