Sexually dimorphic mandibular morphology in the first few years of life

Sex differences in the youngest skeletons are very subtle, and any method that can separate males and females significantly better than chance will be of value. Compounding the problem is a paucity of immature skeletons of documented age and sex. In 1992, S.R.L. examined 62 juvenile mandibles of white and black South Africans of known age and sex (from birth to 19 years) from the Dart Collection to determine if the sexes could be differentiated by morphologic traits. By age 6 years, adult chin shapes were already recognizable. Prior to that age, differences were observed in the shape of the inferior border of the symphysis and outline of the body. The male chin base extends steeply downward relative to the adjacent body, coming to a point or squaring off at the symphysis. In females, the symphysis descends gradually to a more rounded base, and even when pointed, the transition is not abrupt. On the outer border of the corpus, the sides diverge sharply to form a \_/ shape from a roughly horizontal anterior region in males, while the female contour is rounded, reflecting the smoothly curved transition from front to sides. These traits were manifest from the eruption of the central incisors until about 4 years of age. The features were tested on all 19 Dart Collection mandibles in that age range. Average accuracy for three different testers was 81%, and males were consistently identified more accurately than females. This new method was then tested on a known sex sample of 11 individuals from 0 to 7 years of age. These included CT scans of 9 French children and the remains of 2 South African black forensic cases. Sexing accuracy was 82% (9/11). The only two missexed cases were both female and over age 6 years. In conclusion, the results of this study indicate that it is possible to determine the sex of very young mandibles. The new sexually dimorphic morphologic configurations introduced here have demonstrated repeatable discrimination with the highest level of accuracy (81%) reported and tested for this age group. Preliminary research indicates that both the male and female shapes are clearly recognizable in archaeologic and premodern hominids as well as chimpanzees.     

Evolution of Mhc–DRB Introns: Implications for the Origin of Primates

Introns are generally believed to evolve too rapidly and too erratically to be of much use in phylogenetic reconstructions. Few phylogenetically informative intron sequences are available, however, to ascertain the validity of this supposition. In the present study the supposition was tested on the example of the mammalian class II major histocompatibility complex (Mhc) genes of the DRB family. Since the Mhc genes evolve under balancing selection and are believed to recombine or rearrange frequently, the evolution of their introns could be expected to be particularly rapid and subject to scrambling. Sequences of intron 4 and 5 DRB genes were obtained from polymerase chain reaction-amplified fragments of genomic DNA from representatives of six eutherian orders—Primates, Scandentia, Chiroptera, Dermoptera, Lagomorpha, and Insectivora. Although short stretches of the introns have indeed proved to be unalignable, the bulk of the intron sequences from all six orders, spanning >85 million years (my) of evolution, could be aligned and used in a study of the tempo and mode of intron evolution. The analysis has revealed the Mhc introns to evolve at a rate similar to that of other genes and of synonymous sites of non-Mhc genes. No evidence of homogenization or large-scale scrambling of the intron sequences could be found. The Mhc introns apparently evolve largely by point mutations and insertions/deletions. The phylogenetic signals contained in the intron sequences could be used to identify Scandentia as the sister group of Primates, to support the existence of the Archonta superorder, and to confirm the monophyly of the Chiroptera. Received: 26 October 1998 / Accepted: 21 December 1998     

Helifulvanolsäkure—ein neues diterpen mit anomalem kohlenstoffgerüst aus Helichrysum fulvum

The aerial parts of Helichrysum fulvum afforded, in addition to beyerenic acid and ent-kaurenic acid, two new diterpenic acids with the hitherto unknown carbon skeleton of an isotrachylobane type. The structures of these acids, isolated as their methyl esters, were elucidated by extensive NMR studies, some chemical transformations and by X-ray structural analysis of the corresponding acetate. The related alcohol on reaction with pyridinochlorochromate afforded a homoconjugated diene probably formed by fragmentation of a cyclopropyl carbinol. The possible biogenesis of the new carbon skeleton is discussed briefly.     

Jaw transformation with gain of symmetry after Dlx5/Dlx6 inactivation: Mirror of the past?

In modern vertebrates upper and lower jaws are morphologically different. Both develop from the mandibular arch, which is colonized mostly by Hox-free neural crest cells. Here we show that simultaneous inactivation of the murine homeobox genes Dlx5 and Dlx6 results in the transformation of the lower jaw into an upper jaw and in symmetry of the snout. This is the first homeotic-like transformation found in this Hox-free region after gene inactivation. A suggestive parallel comes from the paleontological record, which shows that in primitive vertebrates both jaws are essentially mirror images of each other. Our finding supports the notion that Dlx genes are homeotic genes associated with morphological novelty in the vertebrate lineage.     

Biglycan knockout mice: new models for musculoskeletal diseases

Biglycan is a Class I Small Leucine Rich Proteoglycans (SLRP) that is localized on human chromosome Xq28-ter. The conserved nature of its intron-exon structure and protein coding sequence compared to decorin (another Class I SLRP) indicates the two genes may have arisen from gene duplication. Biglycan contains two chondroitin sulfate glycosaminoglycan (GAG) chains attached near its NH₂ terminus making it different from decorin that has only one GAG chain. To determine the functions of biglycan in vivo, transgenic mice were developed that were deficient in the production of the protein (knockout). These mice acquire diminished bone mass progressively with age. Double tetracycline-calcein labeling revealed that the biglycan deficient mice are defective in their capacity to form bone. Based on this observation, we tested the hypothesis that the osteoporosis-like phenotype is due to defects in cells critical to the process of bone formation. Our data shows that biglycan deficient mice have diminished capacity to produce marrow stromal cells, the bone cell precursors, and that this deficiency increases with age. The cells also have reduced response to tranforming growth factor- (TGF-), reduced collagen synthesis and relatively more apoptosis than cells from normal littermates. In addition, calvaria cells isolated from biglycan deficient mice have reduced expression of late differentiation markers such as bone sialoprotein and osteocalcin and diminished ability to accumulate calcium judged by alizerin red staining. We propose that any one of these defects in osteogenic cells alone, or in combination, could contribute to the osteoporosis observed in the biglycan knockout mice. Other data suggests there is a functional relationship between biglycan and bone morphogenic protein-2/4 (BMP 2/4) action in controlling skeletal cell differentiation. In order to test the hypothesis that functional compensation can occur between SLRPs, we created mice deficient in biglycan and decorin. Decorin deficient mice have normal bone mass while the double biglycan/decorin knockout mice have more severe osteopenia than the single biglycan indicating redundancy in SLRP function in bone tissue. To further determine whether compensation could occur between different classes of SLRPs, mice were generated that are deficient in both biglycan (class I) and fibromodulin, a class II SLRP highly expressed in mineralizing tissue. These doubly deficient mice had an impaired gait, ectopic calcification of tendons and premature osteoarthritis. Transmission electron microscopy analysis showed that like the decorin and biglycan knockouts, they have severely disturbed collagen fibril structures. Biomechanical analysis of the affected tendons showed they were weaker compared to control animals leading to the conclusion that instability of the joints could be the primary cause of all the skeletal defects observed in the fibromodulin/biglycan knockout mice. These studies present important new animal models for musculoskeletal diseases and provide the opportunity to characterize the network of signals that control tissue integrity and function through SLRP activity. Published in 2003.     

G protein-coupled receptor/arrestin3 modulation of the endocytic machinery

Nonvisual arrestins (arr) modulate G protein-coupled receptor (GPCR) desensitization and internalization and bind to both clathrin (CL) and AP-2 components of the endocytic coated pit (CP). This raises the possibility that endocytosis of some GPCRs may be a consequence of arr-induced de novo CP formation. To directly test this hypothesis, we examined the behavior of green fluorescent protein (GFP)-arr3 in live cells expressing beta2-adrenergic receptors and fluorescent CL. After agonist stimulation, the diffuse GFP-arr3 signal rapidly became punctate and colocalized virtually completely with preexisting CP spots, demonstrating that activated complexes accumulate in previously formed CPs rather than nucleating new CP formation. After arr3 recruitment, CP appeared larger: electron microscopy analysis revealed an increase in both CP number and in the occurrence of clustered CPs. Mutant arr3 proteins with impaired binding to CL or AP-2 displayed reduced recruitment to CPs, but were still capable of inducing CP clustering. In contrast, though constitutively present in CPs, the COOH-terminal moiety of arr3, which contains CP binding sites but lacks receptor binding, did not induce CP clustering. Together, these results indicate that recruitment of functional arr3-GPCR complexes to CP is necessary to induce clustering. Latrunculin B or 16 degrees C blocked CP rearrangements without affecting arr3 recruitment to CP. These results and earlier studies suggest that discrete CP zones exist on cell surfaces, each capable of supporting adjacent CPs, and that the cortical actin membrane skeleton is intimately involved with both the maintenance of existing CPs and the generation of new structures.     

Occurrence of articulins and epiplasmins in protists

The cortex of ciliates. dinoflagellates, and euglenoids comprises a unique structure called the epiplasm, implicated in pattern-forming processes of the cell cortex and in maintaining cell shape. Articulins, a novel class of cytoskeletal proteins, are major constituents of the epiplasm in the flagellate Euglena gracilis and the ciliate Pseudomicrothorax dubius. The hallmark of articulins is a core domain of repetitive motifs of alternating valine and proline residues, the VPV-motif. The VPV-motif repeats are 12 residues long. Positively and negatively charged residues segregate in register with valine and proline positions. The VPV-motif is unique to articulins. The terminal domains flanking the core are generally hydrophobic and contain a series of hexa- or heptapeptide repeats rich in glycine and hydrophobic residues. Using molecular and immunological tools we show that articulins are also present in the dinoflagellate Amphidinium carterae and the ciliates Paramecium tetraurelia and Paramecium caudatum, Tetrahymena pyriformis, and Euplotes aediculatus. Our analysis further shows that epiplasmins, a group of epiplasmic proteins first characterized in Paramecium, are also present in all these species. Moreover, we present evidence that epiplasmins and articulins represent two distinct classes of cytoskeletal proteins.     

Novel cytoskeletal proteins in the cortex of Tetrahymena

An important unsolved problem lies in the mechanisms that determine overall size, shape, and the localization of subcellular structures in eukaryotic cells. The membrane skeleton must play a central role in these processes in many cell types, and the ciliate membrane skeleton, or epiplasm, offers favorable opportunities for exploring the molecular determinants of cortical organization. Among the ciliates, Tetrahymena is well suited for the application of a wide range of molecular and cellular approaches. Progress has been made in the identification and sequencing of genes and proteins that encode epiplasmic and cortical proteins. The amino acid sequences of these proteins suggest that they define new classes of cytoskeletal proteins, distinct from the articulin and epiplasmin proteins. We will also discuss recent in vivo and in vitro studies of the regulation of assembly of these cortical proteins. This will include information regarding the down-regulation of epiplasmic proteins during cleavage, their topographic regulation in the cell cycle, and the results of in vitro assembly and binding studies of the epiplasmic C protein.