The Use of Genomic Information for the Conservation of Animal Genetic Diversity

The Saccharomyces cerevisiae leucine-inserting amber suppressor tRNA gene SUP53 (a tRNALeu3 allele) was used to investigate the relationship between precursor tRNA structure and mature tRNA function. This gene encodes a pre-tRNA which contains a 32-base intron. The mature tRNASUP53 contains a 5-methylcytosine modification of the anticodon wobble base. Mutations were made in the SUP53 intron. These mutant genes were transcribed in an S. cerevisiae nuclear extract preparation. In this extract, primary tRNA gene transcripts are end-processed and base modified after addition of cofactors. The base modifications made in vitro were examined, and the mutant pre-tRNAs were analyzed for their ability to serve as substrates for partially purified S. cerevisiae tRNA endonuclease and ligase. Finally, the suppressor function of these mutant tRNA genes was assayed after their integration into the S. cerevisiae genome. Mutant analysis showed that the totally intact precursor tRNA, rather than any specific sequence or structure of the intron, was necessary for efficient nonsense suppression by tRNASUP53. Less efficient suppressor activity correlated with the absence of the 5-methylcytosine modification. Most of the intron-altered precursor tRNAs were successfully spliced in vitro, indicating that modifications are not critical for recognition by the tRNA endonuclease and ligase.

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An In Vitro–In Vivo Simulation Approach for the Prediction of Bioequivalence

Materials ◽

10.3390/ma14030555 ◽

2021 ◽

Vol 14 (3) ◽

pp. 555

Author(s):

Marilena Vlachou ◽

Vangelis Karalis

Keyword(s):

Mathematical Description ◽

Development Process ◽

Bioequivalence Study ◽

In Vitro Dissolution ◽

Monte Carlo Techniques ◽

In Vivo Kinetics ◽

Probability Of Success ◽

Made In

The aim of this study was to develop a new in vitro–in vivo simulation (IVIVS) approach in order to predict the outcome of a bioequivalence study. The predictability of the IVIVS procedure was evaluated through its application in the development process of a new generic product of amlodipine/irbesartan/hydrochlorothiazide. The developed IVIVS methodology is composed of three parts: (a) mathematical description of in vitro dissolution profiles, (b) mathematical description of in vivo kinetics, and (c) development of joint in vitro–in vivo simulations. The entire programming was done in MATLAB® and all created scripts were validated through other software. The IVIVS approach can be implemented for any number of subjects, clinical design, variability and can be repeated for thousands of times using Monte Carlo techniques. The probability of success of each scenario is recorded and finally, an overall assessment is made in order to select the most suitable batch. Alternatively, if the IVIVS shows reduced probability of BE success, the R&D department is advised to reformulate the product. In this study, the IVIVS approach predicted successfully the BE outcome of the three drugs. During the development of generics, the IVIVS approach can save time and expenses.

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Processing of the precursor to a chloroplast ribosomal protein made in the cytosol occurs in two steps, one of which depends on a protein made in the chloroplast

Molecular and Cellular Biology ◽

10.1128/mcb.5.5.1093-1099.1985 ◽

1985 ◽

Vol 5 (5) ◽

pp. 1093-1099

Author(s):

R J Schmidt ◽

N W Gillham ◽

J E Boynton

Keyword(s):

Molecular Weight ◽

Protein Synthesis ◽

Ribosomal Protein ◽

Apparent Molecular Weight ◽

Protein L ◽

Mature Form ◽

Chloroplast Protein Synthesis ◽

Made In

In pulse-chase experiments in which log-phase cells of Chlamydomonas reinhardtii were labeled in vivo for 5 min with H2(35)SO4, fluorographs of immunoprecipitates from whole cell extracts revealed that chloroplast ribosomal proteins L-2, L-6, L-21, and L-29, which are made in the cytosol and imported, appeared in their mature forms. However, in the case of chloroplast ribosomal protein L-18, which is also made in the cytoplasm and imported, a prominent precursor with an apparent molecular weight of 17,000 was found at the end of a 5-min pulse. This precursor was processed to its mature size (apparent molecular weight of 15,500) within the first 5 min of the subsequent chase. As determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the precursor to L-18 formed in vivo was 1.5 kilodaltons smaller than the primary product detected in translations of Chlamydomonas polyadenylated RNA in vitro. Upon a 10-min incubation with a postribosomal supernatant from Chlamydomonas, the 18,500-dalton precursor detected in vitro could be partially converted into a polypeptide that comigrated with the 17,000-dalton precursor detected in extracts of cells labeled in vivo. Under conditions in which the total amounts of chloroplast proteins had been reduced and cells were made to synthesize ribosomes rapidly, the apparent half-life of the 17,000-dalton precursor was extended over that seen in log-phase cells. When chloroplast protein synthesis was inhibited with lincomycin for 3 h before labeling under these conditions, the 17,000-dalton L-18 precursor but not the mature form was found, and the precursor was slowly degraded during a 60-min chase. When cells were placed in the dark for 3 h before labeling, processing of this precursor to the mature form appeared unaffected, but the chloroplast-synthesized ribosomal protein L-26 was detected, indicating that chloroplast protein synthesis was still occurring. We interpret these results to indicate that the maturation of protein L-18 in vivo involves at least two processing steps, one of which depends on a protein made on chloroplast ribosomes.

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Hemodynamics in the abdominal aorta: a comparison of in vitro and in vivo measurements

Journal of Applied Physiology ◽

10.1152/jappl.1994.76.4.1520 ◽

1994 ◽

Vol 76 (4) ◽

pp. 1520-1527 ◽

Cited By ~ 65

Author(s):

J. E. Moore ◽

S. E. Maier ◽

D. N. Ku ◽

P. Boesiger

Keyword(s):

Abdominal Aorta ◽

Velocity Profiles ◽

Flow Reversal ◽

Infrarenal Aorta ◽

Complex Flow ◽

Flow Measurements ◽

In Vivo Measurements ◽

Made In

In vivo measurements of blood velocity profiles are difficult to obtain and interpret, since the parameters that govern the normally highly complex flow situation may not be fully quantified or understood at the time of measurement. In vitro flow models have been used often to better understand vascular hemodynamics. The assumptions made in the design of these models limit the applicability of the results. In this study, in vitro flow measurements made in a carefully designed model of the abdominal aorta were compared with in vivo measurements obtained with magnetic resonance imaging. In the suprarenal aorta, the velocity profiles were mostly forward and axisymmetric in both the in vitro and in vivo cases. In the infrarenal aorta, there was extensive flow reversal noted near the posterior wall in both cases. In the aortic bifurcation, two peaks of flow reversal were noted near the lateral posterior walls, and M-shaped velocity profiles were observed in late diastole. The in vitro and in vivo measurements exhibited good qualitative agreement. The in vitro model was accurate in modeling the in vivo hemodynamics of the abdominal aorta. The complex phenomena observed in vivo were explained on the basis of knowledge gained from the in vitro study.

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The research significance of concomitant use of CAR-CD138-NK and CAR-CD19-NK to target multiple myelomas

European Journal of Inflammation ◽

10.1177/2058739218788968 ◽

2018 ◽

Vol 16 ◽

pp. 205873921878896 ◽

Cited By ~ 3

Author(s):

Songbo Zhao ◽

Zhichao Han ◽

Cheng Ji ◽

Gangli An ◽

Huimin Meng ◽

...

Keyword(s):

Nk Cells ◽

Lymphoblastic Leukemia ◽

Small Subset ◽

Novel Drugs ◽

Prevalent Disease ◽

Cell Acute Lymphoblastic Leukemia ◽

Multiple Myelomas ◽

Made In

Multiple myeloma (MM) is a type of cancer characterized by abnormal proliferation of clonal cells; it is the very dangerous and highly prevalent disease. Although significant progress has been made in clinical research, especially with novel drugs such as bortezomib, lenalidomide, and carfilzomib, most of the patients with MM still suffer from often fetal relapses due to drug resistance. In this study, we aimed to develop immune cells that could specifically target and destroy MM cells. Chimeric antigen receptor–modified NK-92 (CAR-NK92) cells have been very effective against B-cell acute lymphoblastic leukemia (B-ALL); as MM shows high expression of CD138, we constructed CD138-directed CAR-NK-92MI cells (CAR-CD138). It 2is reported that there is a small subset of CD138–/CD19+ MM cells showing, to some extent, stem cell qualities. We therefore generated the CD19-directed CAR-NK-92MI cells (CAR-CD19) as well. These two CAR-NK cells showed strong in vitro biological activity in specifically killing target tumor cells. Thus, the concomitant use of these CAR-NK cells may achieve excellent results in vivo.

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Genetic analysis of silver-fir populations in the Beskids

Acta Societatis Botanicorum Poloniae ◽

10.5586/asbp.2003.015 ◽

2011 ◽

Vol 72 (2) ◽

pp. 115-119 ◽

Cited By ~ 3

Author(s):

Leon Mejnartowicz

Keyword(s):

Genetic Diversity ◽

Genetic Variation ◽

Genetic Analysis ◽

Abies Alba ◽

Genetic Distances ◽

Silver Fir ◽

Geographical Distance ◽

Polymorphic Loci ◽

Expected Heterozygosity ◽

Rare Alleles

Twenty-eight isozymic loci were studied in the Beskid Mts., in four populations of common silver-fir (Abies alba): one in Beskid Makowski (BM) and three populations in Beskid Sądecki (BS). Their genetic variation and diversity were analyzed, and Nei's genetic distances between the populations were calculated. The results show that the geographical distance between the BM population and the three BS populations is reflected in genetic distances. The BM population is clearly distinct from the others. It has the lowest genetic diversity (I = 0.42), percentage of polymorphic loci (%PoL = 64.29) and number of rare alleles (NoRa = 5). Besides, the BM population has the highest observed heterozygosity (Ho = 0.291), which exceeds the expected heterozygosity (He = 0.254), estimated on the basis of the Hardy-Weinberg Principle. On the contrary, BS populations are in the state of equilibrium, which is manifested, in similar values of He = 0.262 and Ho = 0.264.

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Kinematic models of the buccal mass of Aplysia californica.

Journal of Experimental Biology ◽

10.1242/jeb.201.10.1563 ◽

1998 ◽

Vol 201 (10) ◽

pp. 1563-1583 ◽

Cited By ~ 4

Author(s):

R F Drushel ◽

D M Neustadter ◽

I Hurwitz ◽

P E Crago ◽

H J Chiel

Keyword(s):

Neural Control ◽

Kinematic Model ◽

Aplysia Californica ◽

Relative Location ◽

Marine Mollusc ◽

Buccal Mass ◽

Kinematic Models ◽

Made In

The feeding behavior of the marine mollusc Aplysia californica is an intensively studied model system for understanding the neural control of behavior. Feeding movements are generated by contractions of the muscles of the buccal mass. These muscles are internal and cannot be visualized during behavior. In order to infer the movements of the muscles of the buccal mass, two kinematic models were constructed. The first kinematic model assumed that the complex consisting of the pincer-like radula and the underlying odontophore was spherical in shape. In this model, the radula/odontophore was moved anteriorly or posteriorly and the more superficial buccal muscles (I1/I3 and I2) were fitted around it. Although the overall buccal mass shapes predicted by this model were similar to those observed in vivo during protraction, the shapes predicted during retraction were very different. We therefore constructed a second kinematic model in which the shape of the radula/odontophore was based on the shapes assumed by those structures in vitro when they were passively forced into protraction, rest or retraction positions. As each of these shapes was rotated, the second kinematic model generated overall shapes of the buccal mass that were similar to those observed in vivo during swallowing and tearing, and made predictions about the antero-posterior length of the buccal mass and the relative location of the lateral groove. These predictions were consistent with observations made in vivo and in vitro. The kinematic patterns of intrinsic buccal muscles I1 and I2 in vivo were estimated using the second model. Both models make testable predictions with regard to the functions and neural control of intrinsic buccal muscles I2 and I3.

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Targeting Wnt/β-Catenin Pathway for Developing Therapies for Hair Loss

International Journal of Molecular Sciences ◽

10.3390/ijms21144915 ◽

2020 ◽

Vol 21 (14) ◽

pp. 4915 ◽

Cited By ~ 1

Author(s):

Bu Young Choi

Keyword(s):

Hair Loss ◽

Hair Growth ◽

In Vivo Studies ◽

Intracellular Targets ◽

Remarkable Progress ◽

Made In

Persistent hair loss is a major cause of psychological distress and compromised quality of life in millions of people worldwide. Remarkable progress has been made in understanding the molecular basis of hair loss and identifying valid intracellular targets for designing effective therapies for hair loss treatment. Whereas a variety of growth factors and signaling pathways have been implicated in hair cycling process, the activation of Wnt/β-catenin signaling plays a central role in hair follicle regeneration. Several plant-derived chemicals have been reported to promote hair growth by activating Wnt/β-catenin signaling in various in vitro and in vivo studies. This mini-review sheds light on the role of Wnt/β-catenin in promoting hair growth and the current progress in designing hair loss therapies by targeting this signaling pathway.

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In Vivo Titration of Mitomycin C Action by Four Escherichia coli Genomic Regions on Multicopy Plasmids

Journal of Bacteriology ◽

10.1128/jb.183.7.2259-2264.2001 ◽

2001 ◽

Vol 183 (7) ◽

pp. 2259-2264 ◽

Cited By ~ 26

Author(s):

Yan Wei ◽

Amy C. Vollmer ◽

Robert A. LaRossa

Keyword(s):

Escherichia Coli ◽

Mitomycin C ◽

Transcriptional Activation ◽

Efflux Pump ◽

Wild Type ◽

Potent Inducer ◽

E Coli ◽

Genomic Regions

ABSTRACT Mitomycin C (MMC), a DNA-damaging agent, is a potent inducer of the bacterial SOS response; surprisingly, it has not been used to select resistant mutants from wild-type Escherichia coli. MMC resistance is caused by the presence of any of four distinctE. coli genes (mdfA, gyrl, rob, andsdiA) on high-copy-number vectors. mdfAencodes a membrane efflux pump whose overexpression results in broad-spectrum chemical resistance. The gyrI (also called sbmC) gene product inhibits DNA gyrase activity in vitro, while the rob protein appears to function in transcriptional activation of efflux pumps. SdiA is a transcriptional activator of ftsQAZ genes involved in cell division.

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Processing of the precursor to a chloroplast ribosomal protein made in the cytosol occurs in two steps, one of which depends on a protein made in the chloroplast.

Molecular and Cellular Biology ◽

10.1128/mcb.5.5.1093 ◽

1985 ◽

Vol 5 (5) ◽

pp. 1093-1099 ◽

Cited By ~ 27

Author(s):

R J Schmidt ◽

N W Gillham ◽

J E Boynton

Keyword(s):

Molecular Weight ◽

Protein Synthesis ◽

Ribosomal Protein ◽

Apparent Molecular Weight ◽

Protein L ◽

Mature Form ◽

Chloroplast Protein Synthesis ◽

Made In

In pulse-chase experiments in which log-phase cells of Chlamydomonas reinhardtii were labeled in vivo for 5 min with H2(35)SO4, fluorographs of immunoprecipitates from whole cell extracts revealed that chloroplast ribosomal proteins L-2, L-6, L-21, and L-29, which are made in the cytosol and imported, appeared in their mature forms. However, in the case of chloroplast ribosomal protein L-18, which is also made in the cytoplasm and imported, a prominent precursor with an apparent molecular weight of 17,000 was found at the end of a 5-min pulse. This precursor was processed to its mature size (apparent molecular weight of 15,500) within the first 5 min of the subsequent chase. As determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the precursor to L-18 formed in vivo was 1.5 kilodaltons smaller than the primary product detected in translations of Chlamydomonas polyadenylated RNA in vitro. Upon a 10-min incubation with a postribosomal supernatant from Chlamydomonas, the 18,500-dalton precursor detected in vitro could be partially converted into a polypeptide that comigrated with the 17,000-dalton precursor detected in extracts of cells labeled in vivo. Under conditions in which the total amounts of chloroplast proteins had been reduced and cells were made to synthesize ribosomes rapidly, the apparent half-life of the 17,000-dalton precursor was extended over that seen in log-phase cells. When chloroplast protein synthesis was inhibited with lincomycin for 3 h before labeling under these conditions, the 17,000-dalton L-18 precursor but not the mature form was found, and the precursor was slowly degraded during a 60-min chase. When cells were placed in the dark for 3 h before labeling, processing of this precursor to the mature form appeared unaffected, but the chloroplast-synthesized ribosomal protein L-26 was detected, indicating that chloroplast protein synthesis was still occurring. We interpret these results to indicate that the maturation of protein L-18 in vivo involves at least two processing steps, one of which depends on a protein made on chloroplast ribosomes.

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