A cDNA library has been constructed from an RNA preparation highly enriched in silkmoth chorion mRNAs. Many distinct clones have been identified from this library using a stepwise procedure: scoring for infrequent hexanucleotide restriction enzyme recognition sequences; detailed characterization with restriction enzymes that recognize relatively frequent tetranucleotide sequences; probing the arrangement of the corresponding sequences in chromosomal DNA by the Southern procedure; and detailed cross-hybridization analysis. Unique clones, as well as two classes of distinct but related clones, were revealed by hybridization. The cross-hybridization analysis was greatly facilitated by a newly developed, semiquantitative dot hybridization procedure. The same procedure made it feasible to conveniently estimate the relative abundance of several different sequences in an mRNA mixture. Cloned sequences which scored as relatively abundant in total chorion mRNA were tested with stage-specific chorion mRNA at a very stringent criterion of hybridization. They were thus characterized as early, middle or late sequences with respect to development. The characterized cDNA clones can now be used as probes for studying the evolution, chromosomal organization and regulated developmental expression of the chorion multigene families.
The transposable element hobo can be mobilized to induce a variety of genetic abnormalities within the germ‐line of Drosophila melanogaster. Strains containing hobos have 3.0 kb elements and numerous smaller derivatives of the element. By analogy with other transposable element systems, it is likely that only the 3.0 kb elements are capable of inducing hobo mobilization. Here, we report that a cloned 3.0 kb hobo, called HFL1, is able to mediate germ‐line transformation and therefore is an autonomous (fully‐functional) transposable element. Germ‐line transformation was observed when HFL1 and a marked hobo element were co‐injected into recipient embryos devoid of endogenous hobos. Integration did not occur in the absence of the 3.0 kb element. A single copy of the marked hobo transposon inserted at each site, and the target sites were widely distributed throughout the genome. Integration occurred at (or very near) the termini of hobo, without internal rearrangement of the hobo or marker gene sequences. The hobo transformation system will allow us to determine the structural and regulatory features of hobo responsible for its mobilization and will provide novel approaches for the molecular and genetic manipulation of the Drosophila genome.
OBJECTIVE-Lipin, a novel molecular protein expressed by adipocytes, has marked effects on adipose tissue mass, insulin sensitivity, and glucose homeostasis. Thus, we hypothesized that genetic variants within LPIN1 are associated with traits of the metabolic syndrome.RESEARCH DESIGN AND METHODS-A total of 15 single nucleotide polymorphisms (SNPs) covering the LPIN1 gene region were genotyped in an age-and sex-stratified sample of the general population (Monitoring Trends and Determinants on Cardiovascular Diseases Study Augsburg; DNA and phenotypes of 1,416 Caucasians). Ten SNPs were also genotyped for replication in an independent sample of 1,030 subjects recruited throughout Germany. The metabolic syndrome was defined via the sum of its core components and, additionally, by a factor score derived from factor analysis. Permutation-based methods were used to test the association between genetic LPIN1 variants and metabolic traits for empirical significance.RESULTS-Linkage disequilibrium (LD) analysis revealed three LD blocks encompassing LPIN1. We identified three associated three-marker haplotypes: one common haplotype (26.8% frequency) increases the risk for the metabolic syndrome (odds ratio 1.6 [95% CI 1.2-2.2]), while the other two, being less common (5.7 and 4.0%), are strongly associated with lower blood pressure levels (systolic blood pressure 127 Ϯ 18 vs. 135 Ϯ 20 mmHg; P ϭ 0.0001), a lower BMI (24.6 Ϯ 3.6 vs. 26.9 Ϯ 4.1 kg/m 2 ; P ϭ 3.7 ϫ 10 Ϫ7 ) and waist circumference (82 Ϯ 12 vs. 90 Ϯ 12 cm; P ϭ 3.2 ϫ 10 Ϫ8 ), lower A1C levels (5.1 Ϯ 0.7 vs. 5.3 Ϯ 0.9%; P ϭ 0.0002), as well as a lower metabolic syndrome factor score (Ϫ0.67 Ϯ 1.00 vs. 0.04 Ϯ 1.24; P ϭ 1.4 ϫ 10 Ϫ7 ). Furthermore, the frequencies of arterial hypertension (23.7 vs. 46.4%; P ϭ 0.00001), obesity (12.9 vs. 30.8%; P ϭ 0.0003), diabetes (2.2 vs. 8.2%; P ϭ 0.041), and the presence of three or more metabolic syndrome components (3.3 vs. 13.7%; P ϭ 0.002) were significantly lower than in subjects not carrying one of these protective haplotypes.Strong associations were also observed in the replication sample using the same haplotypes but with effects in the opposite direction.CONCLUSIONS-These data suggest that allelic variants of the LPIN1 gene have significant effects in human metabolic traits and thus implicate lipin in the pathophysiology of the metabolic syndrome.
We recently cloned six human importin a proteins that transport specific substrates in complex with importin β into the nucleus. We now compared their absolute expression levels in different human cell lines. We examined their expression regulation during human cell proliferation and differentiation by means of specific antibodies. Proliferation inhibition by starvation of HeLa and HaCaT cells led to a marked decrease in the expression of various nuclear transport factors. In contrast, re-addition of serum increased α-importin expression. We analyzed two models for cell differentiation and found differential importin regulation. Stimulation of rat pancreatic AR42J cell differentiation towards a neuroendocrine phenotype with activin A or towards an acinar phenotype with dexamethasone, caused strong upregulation of importin α3 and α4 expression. Phorbol ester-induced differentiation of human leukemia (HL60) cells towards a macrophage phenotype led to downregulation of importin α1 and α4 expression after 72 hours. Similarly, importins α1 and α4 displayed a strong downregulation when HL60 cells were directed towards a neutrophil phenotype by DMSO treatment. This study is the first to assess all the human importin α isoforms in documenting differential nuclear transport factor regulation during cell proliferation and differentiation.
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