AFLP linkage map of the Japanese quail Coturnix japonica

The quail is a valuable farm and laboratory animal. Yet molecular information about this species remains scarce. We present here the first genetic linkage map of the Japanese quail. This comprehensive map is based solely on amplified fragment length polymorphism (AFLP) markers. These markers were developed and genotyped in an F2 progeny from a cross between two lines of quail differing in stress reactivity. A total of 432 polymorphic AFLP markers were detected with 24 TaqI/EcoRI primer combinations. On average, 18 markers were produced per primer combination. Two hundred and fifty eight of the polymorphic markers were assigned to 39 autosomal linkage groups plus the ZW sex chromosome linkage groups. The linkage groups range from 2 to 28 markers and from 0.0 to 195.5 cM. The AFLP map covers a total length of 1516 cM, with an average genetic distance between two consecutive markers of 7.6 cM. This AFLP map can be enriched with other marker types, especially mapped chicken genes that will enable to link the maps of both species and make use of the powerful comparative mapping approach. This AFLP map of the Japanese quail already provides an efficient tool for quantitative trait loci (QTL) mapping.     

Kinectin Is a Key Effector of RhoG Microtubule-Dependent Cellular Activity

RhoG is a member of the Rho family of GTPases that activates Rac1 and Cdc42 through a microtubule-dependent pathway. To gain understanding of RhoG downstream signaling, we performed a yeast two-hybrid screen from which we identified kinectin, a 156-kDa protein that binds in vitro to conventional kinesin and enhances microtubule-dependent kinesin ATPase activity. We show that RhoG(GTP) specifically interacts with the central domain of kinectin, which also contains a RhoA binding domain in its C terminus. Interaction was confirmed by coprecipitation of kinectin with active RhoG(G12V) in COS-7 cells. RhoG, kinectin, and kinesin colocalize in REF-52 and COS-7 cells, mainly in the endoplasmic reticulum but also in lysosomes. Kinectin distribution in REF-52 cells is modulated according to endogenous RhoG activity. In addition, by using injection of anti-kinectin antibodies that challenge RhoG-kinectin interaction or by blocking anti-kinesin antibodies, we show that RhoG morphogenic activity relies on kinectin interaction and kinesin activity. Finally, kinectin overexpression elicits Rac1- and Cdc42-dependent cytoskeletal effects and switches cells to a RhoA phenotype when RhoG activity is inhibited or microtubules are disrupted. The functional links among RhoG, kinectin, and kinesin are further supported by time-lapse videomicroscopy of COS-7 cells, which showed that the microtubule-dependent lysosomal transport is facilitated by RhoG activation or kinectin overexpression and is severely stemmed upon RhoG inhibition. These data establish that kinectin is a key mediator of microtubule-dependent RhoG activity and suggest that kinectin also mediates RhoG- and RhoA-dependent antagonistic pathways.     

Mapping of FASN and ACACA on two chicken microchromosomes disrupts the human 17q syntenic group well conserved in mammals

Fatty acid synthase and Acetyl-CoA carboxylase are both key enzymes of lipogenesis and may play a crucial role in the weight variability of abdominal adipose tissue in the growing chicken. They are encoded by the FASN and ACACA genes, located on human Chromosome (Chr) 17q25 and on Chr 17q12 or 17q21 respectively, a large region of conserved synteny among mammals. We have localized the homologous chicken genes FASN and ACACA coding for these enzymes, by single-strand conformation polymorphism analysis on different linkage groups of the Compton and East Lansing consensus genetic maps and by FISH on two different chicken microchromosomes. Although synteny is not conserved between these two genes, our results revealed linkage in chicken between FASN and NDPK (nucleoside diphosphate kinase), a homolog to the human NME1 and NME2 genes (non-metastatic cell proteins 1 and 2), both located on human Chr 17q21.3, and also between FASN and H3F3B (H3 histone family 3B), located on human Chr 17q25. The analysis of mapping data from the literature for other chicken and mammalian genes indicates rearrangements have occurred in this region in the mammalian lineage since the mammalian and avian radiation. Received: 8 August 1997 / Accepted: 24 November 1997     

RhoG GTPase Controls a Pathway That Independently Activates Rac1 and Cdc42Hs

RhoG is a member of the Rho family of GTPases that shares 72% and 62% sequence identity with Rac1 and Cdc42Hs, respectively. We have expressed mutant RhoG proteins fused to the green fluorescent protein and analyzed subsequent changes in cell surface morphology and modifications of cytoskeletal structures. In rat and mouse fibroblasts, green fluorescent protein chimera and endogenous RhoG proteins colocalize according to a tubular cytoplasmic pattern, with perinuclear accumulation and local concentration at the plasma membrane. Constitutively active RhoG proteins produce morphological and cytoskeletal changes similar to those elicited by a simultaneous activation of Rac1 and Cdc42Hs, i.e., the formation of ruffles, lamellipodia, filopodia, and partial loss of stress fibers. In addition, RhoG and Cdc42Hs promote the formation of microvilli at the cell apical membrane. RhoG-dependent events are not mediated through a direct interaction with Rac1 and Cdc42Hs targets such as PAK-1, POR1, or WASP proteins but require endogenous Rac1 and Cdc42Hs activities: coexpression of a dominant negative Rac1 impairs membrane ruffling and lamellipodia but not filopodia or microvilli formation. Conversely, coexpression of a dominant negative Cdc42Hs only blocks microvilli and filopodia, but not membrane ruffling and lamellipodia. Microtubule depolymerization upon nocodazole treatment leads to a loss of RhoG protein from the cell periphery associated with a reversal of the RhoG phenotype, whereas PDGF or bradykinin stimulation of nocodazole-treated cells could still promote Rac1- and Cdc42Hs-dependent cytoskeletal reorganization. Therefore, our data demonstrate that RhoG controls a pathway that requires the microtubule network and activates Rac1 and Cdc42Hs independently of their growth factor signaling pathways.     

Iodoamino acid composition of rat thyroglobulin of varying total iodine concentration and fractions isolated by isopycnic ultracentrifugation]

Iodoaminoacid content (iodothyronines, T3 and T4, and iodotyrosines, MIT and DIT) has been determined in enzymatic hydrolysates of thyroglobulin Tg 19S of different iodine content (0.3-0.9%) isolated from equilibrium labeled rats. Preparative equilibrium centrifugation in RbCl density gradients of pure thyroglobulin was used to obtain protein fractions of largely different iodine content (0.2-1.2% I). Thin layer chromatography of total hydrolysates demonstrated that the distribution of iodoaminoacids depends on the total iodine content of each fraction. It is concluded, in agreement with previous results, that the native structure of Tg is an important factor in the regulation of hormone biosynthesis and that even at low iodination levels of Tg. T3 and T4 are synthesized.     

A radiation hybrid map of chicken chromosome 15

We have constructed a radiation hybrid (RH) map of chicken chromosome (GGA) 15. This map can be used as a resource to efficiently map genes to this chromosome. The map has been developed using a 6000 rad chicken-hamster whole-genome radiation hybrid panel (ChickRH6). In total, six microsatellite loci, 18 sequence tagged sites (STSs) from BAC end sequences and 11 genes were typed on the panel. The initial framework map comprised eight markers, and an additional 23 markers were then added to generate the final map. The total map length was 334 centiRay6000 (cR6000). The estimated retention frequency for the data set was 18%. Using an estimated physical length of 21 Mb, the ratio between cR6000 and physical distance over GGA15 was estimated to be 0.063 Mb/cR6000. The present map increases the marker density and the marker resolution on GGA15 and enables fast mapping of new chicken genes homologous to genes from human chromosomes 12 and 22.     

Juvenile social experience affects pairing success at adulthood: congruence with the loser effect?

Social interactions with adults are often critical for the development of mating behaviours. However, the potential role of other primary social partners such as juvenile counterparts is rarely considered. Most interestingly, it is not known whether interactions with juvenile females improve males’ courtship and whether, similar to the winner and loser effects in a fighting context—outcome of these interactions shapes males’ behaviour in future encounters. We investigated the combined effects of male quality and juvenile social experience on pairing success at adulthood in zebra finches (Taeniopygia guttata). We manipulated brood size to alter male quality and then placed males in either same- or mixed-sex juvenile dyads until adulthood. We found that males from reduced broods obtained more copulations and males from mixed-sex dyads had more complete courtships. Furthermore, independent of their quality, males that failed to pair with juvenile females, but not juvenile males, had a lower pairing success at adulthood. Our study shows that negative social experience with peers during adolescence may be a potent determinant of pairing success that can override the effects of early environmental conditions on male attractiveness and thereby supports the occurrence of an analogous process to the loser effect in a mating context.     

Genome wide SNP discovery,analysis and evaluation in mallard (<Emphasis Type="Italic">Anas platyrhynchos</Emphasis>)

Background

Next generation sequencing technologies allow to obtain at low cost the genomic sequence information that currently lacks for most economically and ecologically important organisms. For the mallard duck genomic data is limited. The mallard is, besides a species of large agricultural and societal importance, also the focal species when it comes to long distance dispersal of Avian Influenza. For large scale identification of SNPs we performed Illumina sequencing of wild mallard DNA and compared our data with ongoing genome and EST sequencing of domesticated conspecifics. This is the first study of its kind for waterfowl.

Results

More than one billion base pairs of sequence information were generated resulting in a 16× coverage of a reduced representation library of the mallard genome. Sequence reads were aligned to a draft domesticated duck reference genome and allowed for the detection of over 122,000 SNPs within our mallard sequence dataset. In addition, almost 62,000 nucleotide positions on the domesticated duck reference showed a different nucleotide compared to wild mallard. Approximately 20,000 SNPs identified within our data were shared with SNPs identified in the sequenced domestic duck or in EST sequencing projects. The shared SNPs were considered to be highly reliable and were used to benchmark non-shared SNPs for quality. Genotyping of a representative sample of 364 SNPs resulted in a SNP conversion rate of 99.7%. The correlation of the minor allele count and observed minor allele frequency in the SNP discovery pool was 0.72.

Conclusion

We identified almost 150,000 SNPs in wild mallards that will likely yield good results in genotyping. Of these, ~101,000 SNPs were detected within our wild mallard sequences and ~49,000 were detected between wild and domesticated duck data. In the ~101,000 SNPs we found a subset of ~20,000 SNPs shared between wild mallards and the sequenced domesticated duck suggesting a low genetic divergence. Comparison of quality metrics between the total SNP set (122,000 + 62,000 = 184,000 SNPs) and the validated subset shows similar characteristics for both sets. This indicates that we have detected a large amount (~150,000) of accurately inferred mallard SNPs, which will benefit bird evolutionary studies, ecological studies (e.g. disentangling migratory connectivity) and industrial breeding programs.