Beneficial effects of postnatal skin-to-skin contact.

UNLABELLED: In a randomized trial, Bystrova et al. examined the traditional neonatal practice of swaddling. They demonstrated that extended skin-to-skin contact with the mother is the most effective way to maintain the infant's temperature and decrease the "stress of being born". CONCLUSION: Only through research such as that reported by Bystrova et al. can the advantages or disadvantages of routinely accepted perinatal practices be evaluated.

cDNA cloning, chromosomal localization and evolutionary analysis of mouse vacuolar ATPase subunit D, Atp6m.

The multi-subunit vacuolar ATPase pump uses ATP hydrolysis to move protons into membrane bound compartments. The pump is involved in a variety of cellular functions, including regulation of cytosolic pH, vesicular transport, endocytosis, secretion, and apoptosis. Here, we describe the cDNA cloning and chromosomal mapping of subunit D of murine V-ATPase. The mouse gene, designated Atp6m, maps to Chromosome 12, in a region of high homology with human chromosome 14q24. Evolutionary analysis of subunit D orthologs in a variety of other species reveals that this is a highly conserved protein that has been under remarkably strong negative selection during evolution, most likely reflecting its critical role in multiple cellular processes.

Association between variant plasmid formation and senescence in retroplasmid-containing strains of Neurospora spp.

Serial transfer of Neurospora strains harboring the Mauriceville and Varkud mitochondrial retroplasmids frequently displays erratic growth and senescence. Growth impairment is associated with the formation of variant forms of the retroplasmids that can integrate into the mitochondrial genome, resulting in mtDNA rearrangements and eventual loss of respiratory function. Here, we evaluate the rate at which variant plasmids arise in subcultures of the Mauriceville strain of N. crassa and their association with the senescent phenotype. Although variant plasmid formation preceded senescence, subcultures were found to tolerate variant plasmids for variable lengths of time and no correlation could be made between the specific sequence inserted in the plasmids and the rate or frequency of senescence. In addition, many cultures were found to contain more than one variant plasmid. The lack of concordance between the timing of variant plasmid formation and growth cessation distinguishes these two events, and provides additional insight into the etiology of senescence. We also detected differences in the frequency of senescence between retroplasmid-containing strains of N. crassa and N. intermedia and report the isolation of a strain in which senescence occurs in the absence of variant plasmid formation or detectable alterations in mtDNA. Our findings indicate there are multiple pathways that lead to senescence and suggest there are host-specific mechanisms that suppress the deleterious effects of the variant plasmids.

Dendritic cell-specific MHC class II transactivator contains a caspase recruitment domain that confers potent transactivation activity.

The MHC class II transactivator (CIITA) is a critical transcription factor that regulates genes involved in antigen presentation function. At least three functional forms of CIITA gene products are transcribed from three different promoters. The CIITA gene expressed in dendritic cells (DC-CIITA) has a unique first exon encoding an extended N-terminal region of CIITA. Here, we show that the N terminus of DC-CIITA has high homology to a caspase recruitment domain (CARD) found in components of apoptosis and nuclear factor-kappaB signaling pathways. However, DC-CIITA does not regulate cell death, nor does it induce nuclear factor-kappaB activity. Instead, DC-CIITA is transcriptionally a more potent activator of the MHC class II gene than the form expressed in B cells. A single amino acid substitution in the CARD of DC-CIITA, predicted to disrupt CARD-CARD interactions, diminished the transactivation potential of DC-CIITA. These results indicate that the CARD in the context of CIITA serves as a regulatory domain for transcriptional activity and may function to selectively enhance MHC class II gene expression in dendritic cells.

Identification of members of the Wnt signaling pathway in the embryonic pituitary gland.

Prop1 is one of several transcription factors important for the development of the pituitary gland. Downstream targets of PROP1 and other critical pituitary transcription factors remain largely unknown. We have generated a partial expression profile of the developing pituitary gland containing over 350 transcripts, using cDNA subtractive hybridization between Prop1(df/df) and wild-type embryonic pituitary gland primordia. Numerous classes of genes including transcription factors, membrane associated molecules, and cell cycle regulators were identified in this study. Of the transcripts, 34% do not have sequence similarity to known genes, but are similar to ESTs, and 4% represent novel sequences. Pituitary gland expression of a number of clones was verified using in situ hybridization. Several members of the Wnt signaling pathway were identified in the developing pituitary gland. The frizzled2 receptor, Apc, beta-catenin, groucho, and a novel isoform of TCF4 (officially named Tcf7l2) were identified in developing pituitary libraries. Three N-terminal alternatively spliced Tcf7l2 isoforms are reported here, each of which lacks a DNA-binding domain. Functional studies indicate that these isoforms can act as endogenous inhibitors of Wnt signaling in some contexts. This is the first report of Tcf7l2 and Fzd2 expression in the developing pituitary. These molecules may be important in mediating Wnt signaling during pituitary ontogeny. We expect other transcripts from these libraries to be involved in pituitary gland development.

Senescence associated with the over-replication of a mitochondrial retroplasmid in Neurospora crassa.

Mitochondrial DNA rearrangements and deletions are a prevailing feature of filamentous fungal cultures that undergo senescence. In Neurospora spp., strains containing the Mauriceville and Varkud mitochondrial retroplasmids routinely senesce at elevated temperatures, a process that is initiated by the integration of variant forms of the plasmids into the mitochondrial genome. Here, we describe a strain that is phenotypically distinguishable from previously characterized senescent strains and show that senescence can occur in the absence of plasmid integration and associated alterations in mitochondrial DNA. The MS4416 strain contains a unique variant of the Mauriceville retroplasmid, and undergoes senescence at highly predictable frequencies at 37 degrees, 25 degrees and 18 degrees C. Decline in vegetative growth rate correlates with increased levels of the variant plasmid and alterations in the synthesis of mitochondrially encoded proteins, suggesting that plasmid over-replication interferes with mitochondrial translation. We also report the isolation of a mutant strain that escapes senescence yet still maintains high levels of the variant plasmid. Its ability to tolerate a growth-suppressive retroplasmid suggests that there are mechanisms in Neurospora which compensate for the deleterious effects that plasmid over-replication has on mitochondrial function.

Authoritative knowledge, evidence-based medicine, and behavioral pediatrics.

Evidence-based medicine is the conscientious and judicious use of current best knowledge in making decisions about the care of individual patients, often from well-designed, randomized, controlled trials. Authoritative medicine is the traditional approach to learning and practicing medicine, but no one authority has comprehensive scientific knowledge. Archie Cochrane proposed that every medical specialty should compile a list of all of the randomized, controlled trials within its field to be available for those who wish to know what treatments are effective. This was done first for obstetrics by a group collecting and critically analyzing all of the randomized trials and then indicating procedures every mother should have and those that no mother should have. Support during labor was used as an example. Similar groups are now active in almost all specialties, with information available on the Internet in the Cochrane Database of Systematic Reviews. Developmental-behavioral pediatrics should be part of this movement to evidence-based medicine.

Linear mitochondrial plasmids of F. oxysporum are novel, telomere-like retroelements.

Diverse types of linear RNA and DNA autonomously replicating genetic elements exist in prokaryotic and eukaryotic hosts, yet linear elements that replicate by reverse transcription have not been identified. Here, we report the sequence and organization of two linear mitochondrial plasmids of the fungal plant pathogen F. oxysporum and the characterization of a plasmid-associated reverse transcriptase activity. Plasmids pFOXC2 and pFOXC3 are 1.9 kb in length and have a "clothespin" genomic structure, which includes a terminal hairpin and a telomere-like iteration of a 5 bp sequence at the other terminus. The retroplasmid replication cycle involves novel strategies for copying terminal sequences, which may provide clues concerning the origin of telomerase as well as the evolution of linear DNAs.

The doula: an essential ingredient of childbirth rediscovered.

Eleven randomized control trials examined whether additional support by a trained lay person (called a doula), student midwife or midwife, who provides continuous support consisting of praise, encouragement, reassurance, comfort measures, physical contact and explanations about progress during labor, will affect obstetrical and neonatal outcomes. The women were healthy primigravidas at term. Meta-analysis of these studies showed a reduction in the duration of labor, the use of medications for pain relief, operative vaginal delivery, and in many studies a reduction in caesarian deliveries. At 6 weeks after delivery in one study a greater proportion of doula-supported women were breastfeeding, reported greater self-esteem, less depression, a higher regard for their babies and their ability to care for them compared to the control mothers. Observations during labor showed that fathers remained farther away from mothers than doulas, talked and touched less. When the doula was present with the couple during labor the father offered more personal support. The father-to-be' s presence during labor and delivery is important to the mother and father, but it is the presence of the doula that results in significant benefits in outcome.

Mobile group II introns of yeast mitochondrial DNA are novel site-specific retroelements.

Group II introns aI1 and aI2 of the yeast mitochondrial COXI gene are mobile elements that encode an intron-specific reverse transcriptase (RT) activity. We show here that the introns of Saccharomyces cerevisiae ID41-6/161 insert site specifically into intronless alleles. The mobility is accompanied by efficient, but highly asymmetric, coconversion of nearby flanking exon sequences. Analysis of mutants shows that the aI2 protein is required for the mobility of both aI1 and aI2. Efficient mobility is dependent on both the RT activity of the aI2-encoded protein and a separate function, a putative DNA endonuclease, that is associated with the Zn2+ finger-like region of the intron reading frame. Surprisingly, there appear to be two mobility modes: the major one involves cDNAs reverse transcribed from unspliced precursor RNA; the minor one, observed in two mutants lacking detectable RT activity, appears to involve DNA level recombination. A cis-dominant splicing-defective mutant of aI2 continues to synthesize cDNAs containing the introns but is completely defective in both mobility modes, indicating that the splicing or the structure of the intron is required. Our results demonstrate that the yeast group II intron aI2 is a retroelement that uses novel mobility mechanisms.

The VS catalytic RNA replicates by reverse transcription as a satellite of a retroplasmid.

The mitochondria of certain natural isolates of Neurospora contain both the Varkud plasmid, which encodes a reverse transcriptase, and a small unrelated RNA (VS RNA) that performs RNA-mediated self-cleavage and ligation reactions. Here, we show that VS RNA is transcribed from a VS plasmid DNA template by the Neurospora mitochondrial RNA polymerase using a promoter located immediately upstream of the RNA self-cleavage site that generates monomeric transcripts. VS RNA is then reverse transcribed by the Varkud plasmid reverse transcriptase to yield a full-length (-) strand cDNA, a predicted replication intermediate. Combined with previous genetic evidence, our results indicate that the VS plasmid replicates by reverse transcription as a satellite of the Varkud plasmid. This mode of replication, unprecedented for a satellite RNA, likely reflects the promiscuity of the Varkud plasmid reverse transcriptase, which does not require a specific primer to initiate cDNA synthesis. Our findings indicate how primitive reverse transcriptases with similar relaxed specificity could have facilitated the evolution of new retroelements.

A mitochondrial retroplasmid integrates into mitochondrial DNA by a novel mechanism involving the synthesis of a hybrid cDNA and homologous recombination.

The Mauriceville and Varkud mitochondrial plasmids of Neurospora spp. are closely related, small circular DNAs that propagate via an RNA intermediate and reverse transcription. Although the plasmids ordinarily replicate autonomously, they can also integrate into mitochondrial DNA (mtDNA), yielding defective mtDNAs that in some cases cause senescence. To investigate the integration mechanism, we analyzed four cases in which the Varkud plasmid integrated into the mitochondrial small rRNA gene, three in wild-type subcultures and one in a senescent mutant. Our analysis suggests that the integrations occurred by the plasmid reverse transcriptase template switching between the plasmid transcript and internal sequences in the mitochondrial small rRNA to yield hybrid cDNAs that circularized and recombined homologously with the mtDNA. The integrated plasmid sequences are transcribed, presumably from the mitochondrial small rRNA promoters, resulting in hybrid RNAs containing the 5' segment of the mitochondrial small rRNA linked head-to-tail to the full-length plasmid transcript. Analysis of additional senescent mutants revealed three cases in which the plasmid used the same mechanism to integrate at other locations in the mtDNA. In these cases, circular variant plasmids that had incorporated a mitochondrial tRNA or tRNA-like sequence by template switching integrated by homologous recombination at the site of the corresponding tRNA or tRNA-like sequence in mtDNA. This simple integration mechanism involving template switching to generate a hybrid cDNA that integrates homologously could have been used by primitive retroelements prior to the acquisition of a specialized integration machinery.