Genetic diagnosis of cytoplasmic male sterile cybrid plants of rice

Twelve Japanese rice cultivars were converted to CMS by asymmetric protoplast fusion with MTC-5A, the cytoplasm of which was derived from an indica rice, Chinsurah Boro II. With the exception of the cybrids that had a nucleus from Hoshiyutaka, most of these cybrid plants were sterile. The unique sequence downstream from the mitochondrial atp6 of MTC-5A was specifically amplified in the sterile cybrid plants by PCR. All progenies of the cybrid plants carrying this unique sequence were sterile. On the other hand, in some of the sterile cybrid plants in which the unique sequence was not amplified by PCR, fertility was recovered in their progenies. Somaclonal mutation may have caused sterility in these cybrids. Only the cybrid plants that had the unique sequence detected by PCR were CMS. Thus, the CMS plants can be selected rapidly and easily by PCR, at an early stage of plant regeneration. Soon after transplanting the regenerated plants to a green house, fertile cybrids and sterile cybrids produced by somaclonal mutation can be removed. These findings also show that the unique region downstream from atp6 is tightly linked with the CMS phenotype.     

Mechanism of T5-induced DNA polymerase. II. Characterization of the dead-end complex.

We have shown that bacteriophage T5-induced DNA polymerase replicates short primer-templates (400 to 600 nucleotides long) at a rapid rate initially, followed by a slower rate sustained for much longer periods (Das, S. K., and Fujimura, R. K. (1977) J. Biol. Chem. 252, 8700-8707). In order to explain the slower steady rate and the results of polymer-challenge experiments, we conjectured the presence of a "dead-end complex" formed by the enzyme with the primer-template at the end of the primer elongation process. In this communication we present evidence which indicates that the presumed complex shows a first order kinetics of decay with a half-life of 3.5 min at 37 degrees. Energies of activation for the steady phase of synthesis and the decay of the dead-end complex were both found to be about 23 kcal/mol. This indicates that the dissociation of the aforesaid complex might be the rate-limiting step during the steady phase of synthesis. Correlation between the salt-induced reduction in the half-life of the complex and the increase in the steady rate of synthesis is in agreement with the above mentioned possibility.     

Mechanism of T5-induced DNA polymerase. I. Replication of short primer templates.

Bacteriophage T5-induced DNA polymerase shows an initial phase of rapid synthesis, followed by a slower steady rate for much longer periods, with short DNA primer-templates (400 to 600 nucleotides long), in vitro. On extrapolating the line of steady rate back to 0 min, an intercept is obtained on the ordinate. With large DNA primer-templates, such as denatured T5 DNA (average chain length approximately 50,000 bases), the rate of synthesis remains constant and is equal to the initial rate obtained with short primer-templates. The zero time intercept was proportional to the amount of enzyme used and independent of temperature. Polymer challenge experiments indicate that the initial phase of rapid synthesis can be attributed to the processive mode of synthesis by T5 DNA polymerase. After synthesizing a stretch of DNA processively for about 200 nucleotide residues, the enzyme apparently forms a "dead-end complex" with the primer-templates used and must dissociate from the primer-template in order to resume synthesis. The average size of the product made processively, during various phase of synthesis, remains invariant and is in good agreement with the size of the zero time intercept per enzyme molecule.     

Information, discrimination and divergence in cytology. III. Optimization of classification of Papanicolaou smears

The performance of a cytology laboratory can be objectively quantitated as the total discrimination, a defined quantity of information. The total discrimination is dependent on the number of categories used in gynecologic cytology and on the corresponding histologic states; over-classification results in a higher rate of misinformation and reduced total discrimination. Total divergence is another measure of the association between cytologic categories and histologic states; in contrast to the total discrimination, the total divergence does not require a one-to-one correspondence between the cytologic categories and the histologic states. Using data from the Gynecologic Cytology Laboratory of the University of Minnesota, the total discrimination was maximized when gynecologic cytology used three categories of diagnosis, consisting of (1) normal, atypical benign or reactive atypia, (2) cervical intraepithelial neoplasia (CIN) and (3) all malignancies. The use of four categories, (1) normal, atypical benign or reactive atypia, (2) mild or moderate dysplasia, (3) severe dysplasia or squamous carcinoma in situ and (4) all malignancies, was almost equally informative. Observations on the total divergences resulted in similar conclusions. These findings generally support the recommendation of the consensus workshop sponsored by the National Cancer Institute (the Bethesda System nomenclature) to group all degrees of CIN into two large categories.     

Mechanisms of somatic embryogenesis in cell cultures: Physiology,biochemistry, and molecular biology

Summary One of the most characteristic cell functions in plants is totipotency. Somatic embryogenesis can be regarded as a model system for the investigation of mechanisms of totipotency, because a high frequency and synchronous embryogenic system from single somatic cells has been established in carrot suspension cultures. Four phases are recognized in this process, and several molecular markers, viz. polypeptides, mRNAs, antigens against monoclonal antibodies, can be detected during the expression of totipotency, but they disappear during its loss. Four organ-specific genes have been isolated from hypocotyls and roots by differential screening. They were expressed preferentially after the globular-heart stages of embryogenesis, and were strongly suppressed by auxin. A CEM 1 gene was isolated by differential screening of embryogenic cell clusters. This gene was expressed strongly and transiently during the proglobular and globular stages. The sequence of CEM 1 was found to encode a polypeptide showing high homology to the elongation factor isolated from eucaryotic cells. Thus good progress is being made in understanding the basic mechanisms of somatic embryogenesis. Presented in the Session-in-Depth Developmental Biology of Embryogenesis at the 1991 World Congress on Cell and Tissue Culture, Anaheim, California, June 16–20, 1991.     

Inflammatory pseudotumor of the lung diagnosed as granulomatous lesion by preoperative brushing cytology. A case report

The clinical and cytologic features of a case of inflammatory pseudotumor of the lung are presented. Chest roentgenograms revealed a solitary circumscribed round mass in a nine-year-old boy. The mass was diagnosed as a granulomatous lesion by bronchoscopic brushing cytology. Although smears and cultures of sputum and brushing specimens were negative for tuberculosis, a tuberculin reaction was positive and antitubercular therapy was instituted. Since the mass had grown further after six months of therapy, an open lung biopsy was performed to resect the lesion and establish the diagnosis. Imprint smears of the cut surface of the lesion showed cytologic features similar to those of the brushings: short, spindle-shaped cells with a tendency to be arranged in stori-form patterns against a background of minimal necrotic debris. Histopathology established the final diagnosis of inflammatory pseudotumor, a rare granulomatous lesion radiologically resembling a true tumor. Since this lesion usually occurs in younger patients, inflammatory pseudotumor should be considered in pediatric cases with an intrapulmonary lesion that shows histiocytic spindle-shaped cells in stori-form patterns, but whose smears and cultures test negative for tuberculosis.     

Postgenomic futures: translations across the machine-nature border in systems biology

Abstract

This article discusses the production of new “postgenomic” knowledges that aim to be more ecological and “wholistic” than the reductionist genetics of the last forty years. It examines systems biology and, briefly, developmental systems theory, which are two approaches that attempt to model complexities in biology. System biological metaphors and languages have been in part taken from engineering models of automobiles, airplanes and robots and then applied to complex living systems. Systems biology is only the most recent example and perhaps an excellent case in which to study this movement back and forth across the machine-living organism border in Euro-American biology to track how what we know to be nature and machine is constituted. This article argues for a careful analysis of this historical production specifically around the question of what is lost in translation at these border crossings and their potential consequences.     

Noisy Interlimb Coordination Can Be a Main Cause of Freezing of Gait in Patients with Little to No Parkinsonism

Freezing of gait in patients with Parkinson’s disease is associated with several factors, including interlimb incoordination and impaired gait cycle regulation. Gait analysis in patients with Parkinson’s disease is confounded by parkinsonian symptoms such as rigidity. To understand the mechanisms underlying freezing of gait, we compared gait patterns during straight walking between 9 patients with freezing of gait but little to no parkinsonism (freezing patients) and 11 patients with Parkinson’s disease (non-freezing patients). Wireless sensors were used to detect foot contact and toe-off events, and the step phase of each foot contact was calculated by defining one stride cycle of the other leg as 360°. Phase-resetting analysis was performed, whereby the relation between the step phase of one leg and the subsequent phase change in the following step of the other leg was quantified using regression analysis. A small slope of the regression line indicates a forceful correction (phase reset) at every step of the deviation of step phase from the equilibrium phase, usually at around 180°. The slope of this relation was smaller in freezing patients than in non-freezing patients, but the slope exhibited larger step-to-step variability. This indicates that freezing patients executed a forceful but noisy correction of the deviation of step phase, whereas non-freezing patients made a gradual correction of the deviation. Moreover, freezing patients tended to show more variable step phase and stride time than non-freezing patients. Dynamics of a model of two coupled oscillators interacting through a phase resetting mechanism were examined, and indicated that the deterioration of phase reset by noise provoked variability in step phase and stride time. That is, interlimb coordination can affect regulation of the gait cycle. These results suggest that noisy interlimb coordination, which probably caused forceful corrections of step phase deviation, can be a cause of freezing of gait.     

Annexin A4 Is Involved in Proliferation,Chemo-Resistance and Migration and Invasion in Ovarian Clear Cell Adenocarcinoma Cells

Ovarian clear cell adenocarcinoma (CCC) is the second most common subtype of ovarian cancer after high-grade serous adenocarcinomas. CCC tends to develop resistance to the standard platinum-based chemotherapy, and has a poor prognosis when diagnosed in advanced stages. The ANXA4 gene, along with its product, a Ca⁺⁺-binding annexin A4 (ANXA4) protein, has been identified as the CCC signature gene. We reported two subtypes of ANXA4 with different isoelectric points (IEPs) that are upregulated in CCC cell lines. Although several in vitro investigations have shown ANXA4 to be involved in cancer cell proliferation, chemoresistance, and migration, these studies were generally based on its overexpression in cells other than CCC. To elucidate the function of the ANXA4 in CCC cells, we established CCC cell lines whose ANXA4 expressions are stably knocked down. Two parental cells were used: OVTOKO contains almost exclusively an acidic subtype of ANXA4, and OVISE contains predominantly a basic subtype but also a detectable acidic subtype. ANXA4 knockdown (KO) resulted in significant growth retardation and greater sensitivity to carboplatin in OVTOKO cells. ANXA4-KO caused significant loss of migration and invasion capability in OVISE cells, but this effect was not seen in OVTOKO cells. We failed to find the cause of the different IEPs of ANXA4, but confirmed that the two subtypes are found in clinical CCC samples in ratios that vary by patient. Further investigation to clarify the mechanism that produces the subtypes is needed to clarify the function of ANXA4 in CCC, and might allow stratification and improved treatment strategies for patients with CCC.