Development of sterile ovules on bisexual cones of Gnetum gnemon (Gnetaceae).

Angiosperms and Gnetales (Ephedra, Gnetum, Welwitschia) represent the only seed plants that regularly produce bisexual cones. Unfortunately, the fertility and function of ovules formed on bisexual cones of Gnetales have remained unclear. Some reports indicate that the ovules are sterile while others indicate that they may develop into seeds. This study demonstrates three different developmental patterns of ovules formed on bisexual cones of Gnetum gnemon. Type I ovules did not develop at all after pollination and represented the majority of ovules on each cone. Type II ovules enlarged slightly after pollination due to the enlargement of nucellar tissue. Type III ovules were typically found on the terminal whorl and developed into seed-like structures. The enlargement was due to proliferation of megagametophyte tissue. Sectioned material revealed that megagametophytes show altered development compared to those found in functional female ovules. None of the ovules studied contained embryos, and thus all were sterile. Densitometry of 4',6-diamidino-2-phenylindole (DAPI)- stained sections revealed that megagametophyte nuclei formed in the sterile ovules are unreduced (diploid) and thus do not form viable female gametes.

Double Fertilization in Gnetum gnemon: The Relationship between the Cell Cycle and Sexual Reproduction.

Gnetum gnemon, a nonflowering seed plant and member of the Gnetales, expresses a rudimentary pattern of double fertilization that results in the formation of two zygotes per pollen tube. The process of double fertilization in G. gnemon was examined with light and fluorescence microscopy, and the DNA content of various nuclei involved in sexual reproduction was quantified with 4[prime],6-diamidino-2-phenylindole microspectrofluorometry.Male and female gamete nuclei pass through the synthesis phase of the cell cycle and increase their DNA content from 1C to 2C before fertilization. Each of the two zygotes found in association with a pollen tube is diploid and contains the 4C quantity of DNA at inception. Based on these results as well as previous studies of nuclear DNA content in plant sperm, eggs, and zygotes, three fundamental and distinct patterns of gamete karyogamy among seed plants can be circumscribed: (1) G1 karyogamy, in which male and female gametes contain the 1C quantity of DNA throughout karyogamy and the zygote undergoes DNA replication; (2) S-phase karyogamy, in which gamete nuclei initiate fusion at 1C but pass through the S phase of the cell cycle before completely fusing; and (3) G2 karyogamy, in which male and female gamete nuclei pass through the S phase of the cell cycle before the onset of fertilization. Our results show definitively a pattern of G2 karyogamy in G. gnemon.

The competitive inhibition of Trichoderma reesei C30 cellobiohydrolase I by guanidine hydrochloride.

The p-nitrophenylcellobiosidase (PNPCase) activity of Trichoderma reesei cellobiohydrolase I (CBH I) was competitively inhibited by concentrations of guanidine hydrochloride (Gdn HCl) that did not affect the tryptophan fluorescence of this enzyme. The Km of CBH I, 3.6 mM, was increased to 45.4 mM in the presence of 0.14 M Gdn HCl, the concentration that was required to inhibit the enzyme by 50%. A similar concentration of lithium chloride and urea had little effect on the PNPCase activity of CBH I. Maximal inhibition was pH dependent, occurring in the range of pH 4.0 to 5.0, which is in the range for maximal activity. Analysis of the inhibition data indicated that 1.2 molecules of Gdn HCl combined reversibly with 1 molecule of CBH I. Other hydrolases and proteases were also inhibited by Gdn HCl. It is suggested that the inhibition of CBH I by Gdn HCl occurs as a result of the interaction between the positively charged guanidinium group of Gdn HCl and the carboxylate group of glutamic acid 126, postulated to be in the catalytic center of this enzyme.

Comparison of the hydrolytic activity and fluorescence of native, guanidine hydrochloride-treated and renatured cellobiohydrolase I from Trichoderma reesei.

Guanidine hydrochloride (GdnHCl) is an effective agent for the elution of cellulase protein from unhydrolyzed cellulosic residues, but once eluted the enzyme is inactive. The studies described in this paper examine the effect of GdnHCl on the hydrolytic activity and tryptophan fluorescence of cellobiohydrolase I (CBH I) from Trichoderma reesei. CBH I was found to be completely inactivated by 0.25 M GdnHCl, but higher concentrations of GdnHCl were required to partially unfold this enzyme, as determined from the measurement of a decrease in its tryptophan fluorescence. Binding of CBH I to microcrystalline cellulose was prevented by 4 M GdnHCl, suggesting that a conformational change of CBH I resulted in the loss of substrate binding. Removal of the denaturant from CBH I by dialysis or gel filtration allowed the kinetics of the reactivation of CBH I, after 4 M GdnHCl treatment, to be studied. The fluorescence and specific hydrolytic activity of native and renatured CBH I were comparable. It is concluded, therefore, that GdnHCl may be used to elute cellulase components, such as CBH I, adsorbed on undigested cellulosic substrates since this component can easily be renatured and subsequently reused.

Focus on the family as a factor in differential treatment outcome.

The prognostic relationship between prehospital living arrangements and treatment outcome was explored. Drug (N = 17) and alcohol (N = 19) patients who had lived with parents, wives, nonrelatives, or alone before voluntary admission to an inpatient substance abuse program were compared on changes in social dysfunctioning and symptomatology as a result of treatment. Ratings on each patient's level of dysfunctioning and symptomatology were made following intake into the program and within a 3-day period after receiving a regular discharge. While treatment had a positive effect on all groups, multivariate analysis of covariance showed a significant (P less than .009) differential rate of improvement between the pretreatment residential setting groups. Those who had lived alone or with nonrelatives prior to admission changed the most, becoming significantly less dysfunctional and symptomatic. Those coming from parental families changed the least improvement, while those who had lived with their spouses showed moderate improvement. Findings were consistent for both drug and alcohol patients. Results indicate that the type of home environment from which substance abusers come before engaging in treatment significantly influences their receptivity to rehabilitation. Findings suggest the need for assessment of the family environment and utilization of community resources as a means of enhancing adjustment.

The alcoholic's perception of the ward as a predictor of aftercare attendance.

The present study attempted to determine whether attendance in aftercare services could be predicted from the alcoholic's perception of the inpatient ward environment. Thirty-five Ss who completed inpatient treatment and were eligible for outpatient group therapy were followed up 3 months after discharge. One factor of the Ward Atmosphere Scale, autonomy, significantly differentiated attenders (N = 13) from nonattenders (N = 22); attenders perceived more autonomy on the ward than did nonattenders. Two additional factors, aggression and insight, also differentiated the groups, although not to a statistically significant degree. Attenders were found to perceive more encouragement by the staff to express openly their angry feelings and to share their personal selves with others than were nonattenders.