Characterization and sequence analysis of a developmentally regulated putative cell wall protein gene isolated from soybean

A cDNA clone, pTU04, which hybridizes to two different sizes of mRNA on Northern blots was isolated from soybean suspension culture cell poly(A) RNA. Northern analysis reveals that meristematic tissue produces a 1050-nucleotide mRNA while quiescent mature cells produce primarily a 1220-nucleotide mRNA homologous to pTU04. The cDNA and its corresponding genomic clone have been partially characterized. The nucleotide sequence of the gene predicts a proline-rich protein, designated SbPRP1, which contains a signal peptide sequence and 43 repeats of a sequence consisting primarily of Pro-Pro-Val-Tyr-Lys (CCA-CCA-GTT-TAC-AAG). From nuclease S1 and hybrid-select translation analyses, the cDNA clone pTU04 appears to represent the mRNA for the mature tissue 1220-nucleotide RNA observed on Northern blots. Although there is no direct proof that the encoded protein is a cell wall protein, it has the properties similar to previously isolated cell wall proteins: 1) it is very basic with a high content of Pro, Tyr, and Lys; 2) it has similar hydropathic properties; and 3) its repeating unit shares sequence homology with that of more highly characterized cell wall proteins, generally termed extensin (Chen, J., and Varner, J. E. (1985) EMBO J. 4, 2145-2151; Smith, J. J., Muldoon, E. P., Willard, J. J., and Lamport, D. T. A. (1986) Phytochemistry 25, 1021-1030.     

Production of bovine identical twins via transfer of demi-embryos without zonae pellucidae

Embryos were collected nonsurgically from n?turally-cycling or superovulated donors 7 d after estrus. Forty-four morulae, early blastocysts and blastocysts classified as good to excellent were bisected using a fine glass needle to produce forty-four identical demi-embryos. The bisected demi-embryos, without zonae pellucidae, were nonsurgically transferred, either by twin or single transfer. An additional forty-eight embryos were collected from the same donors and transferred as a control. Among the twin transfers, 8 of the 13 recipients became pregnant (61.5%). Seven of them conceived twin fetuses (87%) and one a single fetus. However, only two sets of normal identical twin calves were obtained. Among the single transfers, 72.6% (45/62) of bisected embryos without zonae pellucidae resulted in pregnancy, of which 48.4% (30/62) were identical twins, and 24.2% (15/62) were singletons. Another 27.4% (17/62) of the recipients did not became pregnant. The pregnancy rate for whole embryos with zonae pellucidae was 72.9% (35/48). These data show that there was no significant difference between the pregnancy rates of bisected embryos without zonae pellucidae and whole embryos with zonae pellucidae transferred 7 d after estrus. Bisection of bovine embryos was simplified and even morula stage embryos were transferred without zonae pellucidae.     

Low Mr GTP-binding proteins in human platelets: cyclic AMP-dependent protein kinase phosphorylates m22KG(I) in membrane but not c21KG in cytosol

We have purified and characterized two kinds of GTP-binding proteins with Mr of 22,000 in human platelet membrane (main; m22KG(I), minor; m22KG(II)) (Nagata, K. and Nozawa, Y. (1988) FEBS Lett. 238, 90-94). In this study, the main GTP-binding protein (m22KG(I)) was found to be phosphorylated by cyclic AMP-dependent protein kinase (A-kinase), but not by protein kinase C. About 0.5 mol of phosphate was maximally incorporated into one mol of the protein and this phosphorylation was inhibited in the presence of A-kinase inhibitor. Phosphorylation of m22KG(I) did not alter either its GTP-binding or GTPase activity. When m22KG(I) was incubated alone or in the presence of 100 microM guanosine 5'-(3-O-thio)triphosphate (GTP gamma S) and then exposed to A-kinase, no significant changes in the level of phosphorylation were observed. On the other hand, the most abundant GTP-binding protein with Mr of 21,000 (c21KG) in human platelet cytosol, which was identified as a transformation suppressor gene product (rap 1 protein, smg p21 and Krev-1 protein), was not phosphorylated by A-kinase under the same condition. However, c21KG was phosphorylated by A-kinase after pretreatment with alkaline phosphatase.     

Ontogenetic transition of wound healing pattern in rat skin occurring at the fetal stage

Full-thickness excisional wounds were made in the dorsal skin of rat fetuses at day 16 and day 18 of gestation. A small patch of skin surrounding the open wound was cut out, mounted on a plastic ring and incubated in an organ culture system. In the presence of serum, the open wound in the day-16 fetal skin closed within three days of culture. During the wound-closure process, no new structures were formed in the wound space, and no conspicuous changes were noted in the histological architecture of the surrounding skin during culture, indicating that the wound closure may result from a centripetal movement of the surrounding skin only. In contrast, the size of the open wound in the day-18 fetal skin remained almost unchanged for one week, but a thin acellular network spread over the wound space within one day of culture. The predominant component of the network was cross-linked fibrin, as disclosed by scanning electron microscopy and sodium dodecylsulfate-polyacrylamide gel electrophoresis followed by immunoblotting. The network served as a scaffold for the ingrowth of fibroblast-like cells. These stage-dependent differences in fetal wound healing were consistent with an in vivo study showing that the day-16 wound was covered with the surrounding skin itself, whereas the day-18 wound was covered with newly formed epidermis and invaded by inflammatory cells. The present investigation strongly indicates the prenatal occurrence of a fetal-to-adult transition in the wound-healing pattern of rat skin.     

The effects of guanine and cytosine variation on dinucleotide frequency and amino acid composition in the human genome

Summary One hundred twelve human DNA sequences were analyzed with respect to dinucleotide frequency and amino acid composition. The variation in guanine and cytosine (G+C) content revealed: (1) at 2–3 and 3-1 doublet positions CG discrimination is attenuated at high G+C, but TA disfavor is enhanced, and (2) several amino acids are subject to G+C change. These findings have been reported in part for collections of sequences from various species. The present study confirms that in a single organism-the human-the G+C effects do exist. Aspects of the argument that connects G+C with protein thermal stability are also discussed.