Interpreting genetic effects through models of cardiac electromechanics.

Multiscale models of cardiac electromechanics are being increasingly focused on understanding how genetic variation and environment underpin multiple disease states. In this paper we review the current state of the art in both the development of specific models and the physiological insights they have produced. This growing research body includes the development of models for capturing the effects of changes in function in both single and multiple proteins in both specific expression systems and in vivo contexts. Finally, the potential for using this approach for ultimately predicting phenotypes from genetic sequence information is discussed.

Order-preserving principles underlying genotype-phenotype maps ensure high additive proportions of genetic variance.

In quantitative genetics, the degree of resemblance between parents and offspring is described in terms of the additive variance (V(A)) relative to genetic (V(G)) and phenotypic (V(P)) variance. For populations with extreme allele frequencies, high V(A)/V(G) can be explained without considering properties of the genotype-phenotype (GP) map. We show that randomly generated GP maps in populations with intermediate allele frequencies generate far lower V(A)/V(G) values than empirically observed. The main reason is that order-breaking behaviour is ubiquitous in random GP maps. Rearrangement of genotypic values to introduce order-preservation for one or more loci causes a dramatic increase in V(A)/V(G). This suggests the existence of order-preserving design principles in the regulatory machinery underlying GP maps. We illustrate this feature by showing how the ubiquitously observed monotonicity of dose-response relationships gives much higher V(A)/V(G) values than a unimodal dose-response relationship in simple gene network models.

BAC-based upgrading and physical integration of a genetic SNP map in Atlantic salmon.

A better understanding of the genotype-phenotype correlation of Atlantic salmon is of key importance for a whole range of production, life history and conservation biology issues attached to this species. High-density linkage maps integrated with physical maps and covering the complete genome are needed to identify economically important genes and to study the genome architecture. Linkage maps of moderate density and a physical bacterial artificial chromosome (BAC) fingerprint map for the Atlantic salmon have already been generated. Here, we describe a strategy to combine the linkage mapping with the physical integration of newly identified single nucleotide polymorphisms (SNPs). We resequenced 284 BAC-ends by PCR in 14 individuals and detected 180 putative SNPs. After successful validation of 152 sequence variations, genotyping and genetic mapping were performed in eight salmon families comprising 376 individuals. Among these, 110 SNPs were positioned on a previously constructed linkage map containing SNPs derived from expressed sequence tag (EST) sequences. Tracing the SNP markers back to the BACs enabled the integration of the genetic and physical maps by assigning 73 BAC contigs to Atlantic salmon linkage groups.

Reducing inter-replicate variation in fourier transform infrared spectroscopy by extended multiplicative signal correction.

Fourier transform infrared (FT-IR) spectroscopy is a powerful tool for characterizing biological tissues and organisms, but it is plagued by replicate variation of various sources. Here, a method for estimating and correcting unwanted replicate variation in multivariate measurement signals, based on extended multiplicative signal correction (EMSC), is presented. Systematic patterns of unwanted methodological variations are estimated from replicate spectra, modeled by a linear subspace model, and implemented into EMSC. The method is applied to FT-IR spectra of two different sets of microorganisms (different double gene knockout strains of Saccharomyces cerevisiae and different species of Listeria) and compared to other preprocessing methods used in FT-IR absorption spectroscopy of microorganisms. The EMSC replicate correction turns out to perform best among the compared methods.

Specific developmental gene silencing in the honey bee using a homeobox motif.

Manipulating the expression of genes in species that are not currently used as genetic models will provide comparative insights into the evolution of gene functions. However the experimental tools in doing so are limited in species that have not served as models for genetic studies. We have examined the effects of double stranded RNA (dsRNA) in the honey bee, an insect with considerably basic scientific interest. dsRNA derived from a 300 bp stretch of the E30 homeobox motif was injected into honey bee embryos at the anterior pole in the preblastoderm stage. We found that the dsRNA fragment successfully disrupted the protein expression of the target gene throughout the whole embryo. The disruption caused deficient phenotypes similar to known loss of function mutants of Drosophila engrailed, whereas embryos injected with nonsense dsRNA showed no abnormalities. We show that the large size of the honey bee egg (D: 0.3 mm, L: 1.6 mm) and the long preblastoderm stage (11-12 h) can be exploited to generate embryos with partial disruption of gene function, which may provide an elegant alternative to classical chimeric analyses. This is the first report of targeted disruption of gene function in the honey bee, and the results prove that the chosen target gene is a functional ortholog to engrailed in Drosophila.

Gene regulatory networks generating the phenomena of additivity, dominance and epistasis.

We show how the phenomena of genetic dominance, overdominance, additivity, and epistasis are generic features of simple diploid gene regulatory networks. These regulatory network models are together sufficiently complex to catch most of the suggested molecular mechanisms responsible for generating dominant mutations. These include reduced gene dosage, expression or protein activity (haploinsufficiency), increased gene dosage, ectopic or temporarily altered mRNA expression, increased or constitutive protein activity, and dominant negative effects. As classical genetics regards the phenomenon of dominance to be generated by intralocus interactions, we have studied two one-locus models, one with a negative autoregulatory feedback loop, and one with a positive autoregulatory feedback loop. To include the phenomena of epistasis and downstream regulatory effects, a model of a three-locus signal transduction network is also analyzed. It is found that genetic dominance as well as overdominance may be an intra- as well as interlocus interaction phenomenon. In the latter case the dominance phenomenon is intimately connected to either feedback-mediated epistasis or downstream-mediated epistasis. It appears that in the intra- as well as the interlocus case there is considerable room for additive gene action, which may explain to some degree the predictive power of quantitative genetic theory, with its emphasis on this type of gene action. Furthermore, the results illuminate and reconcile the prevailing explanations of heterosis, and they support the old conjecture that the phenomenon of dominance may have an evolutionary explanation related to life history strategy.

Early developmental processes in the fertilised honeybee (Apis mellifera) oocyte.

The behaviour of sperm from egg penetration until creation of the zygote, the development of the maternal pronucleus, and the two first cleavage divisions were studied by use of fluorescence microscopy. It was found that 4-12 sperm penetrate the egg membranes prior to oviposition. Contrary to previous reports, we found that only 1-7 sperm move from their initial location just beneath the vitelline membrane and into the cytoplasm, where they develop into paternal pronuclei. At the time of oviposition, the oocyte nucleus was usually at the stage of metaphase I, rather than anaphase I as previously reported. At 26+/-2.5 minutes the meiotic process had entered the stage of metaphase II. The paternal and maternal pronuclei formed at 55+/-2.6 minutes, and they fused at 93+/-7.3 minutes. The mitotic division of the zygote was completed at 119+/-6.5 minutes.

Description and analysis of switchlike regulatory networks exemplified by a model of cellular iron homeostasis.

The post-transcriptional regulation of factors involved in the maintenance of cellular iron homeostasis is exerted by iron regulatory proteins (IRPs), which bind to mRNA structures known as iron regulatory elements (IREs). The IRP-IRE interactions are regulated by the intracellular iron level by affecting the binding affinity, synthesis and stability of the IRPs. A model of this homeostasis phenomenon is described and analysed within a methodological framework specifically designed for handling complex systems with steep sigmoidal input/output relationships between the state variables. According to the analysis there is only one threshold regulated homeostatic point. Approximate values for its coordinates, and the conditions ensuring its existence, may be given in terms of parameters. The analysis also provides some tentative insight into the evolution of the regulatory system. A comparison between analytical and numerical estimates of the position of the stationary point as a function of the steepness of the sigmoidal interactions show that the analytical approximations agree quite well with the numerical ones. The results show that we are able to obtain a deeper analytical insight by this methodological framework than what is achievable by most alternative approaches. We find this type of insight to be of considerable heuristic value in connection with the numerical simulation work which normally must be done to unfold the predictive potential of a complex model.

A methodological basis for description and analysis of systems with complex switch-like interactions.

A wide range of complex systems appear to have switch-like interactions, i.e. below (or above) a certain threshold x has no or little influence on y, while above (or below) this threshold the effect of x on y saturates rapidly to a constant level. Switching functions are frequently described by sigmoid functions or combinations of these. Within the context of ordinary differential equations we present a very general methodological basis for designing and analysing models involving complicated switching functions together with any other non-linearities. A procedure to determine position and stability properties of all stationary points lying close to a threshold for one or several variables, so-called singular stationary points, is developed. Such points may represent homeostatic states in models, and are therefore of considerable interest. The analysis provides a profound insight into the generic effects of steep sigmoid interactions on the dynamics around homeostatic points. It leads to qualitative as well as quantitative predictions without using advanced mathematical methods. Thus, it may have an important heuristic function in connection with numerical simulations aimed at unfolding the predictive potential of realistic models.

A mathematical framework for describing and analysing gene regulatory networks.

This paper presents a mathematical framework for describing and analysing gene regulatory networks by autonomous differential equations. It represents an improvement on existing frameworks in that it may handle a wider range of gene regulatory mechanisms. Gene regulatory networks are frequently threshold-dominated, i.e. genes are activated only when the concentration of certain gene products lie between definite thresholds. Here, the concept of regulatory domain is introduced to describe these regions in the phase space. To each regulatory domain is associated an indicator function whose value is 1 inside and 0 outside the domain. The indicator functions thus reflect the logical structure of the network. The sharp borders between the regulatory domains may be smoothed by replacing the logical step functions by continuous sigmoids or so-called logoid functions. A logoid function coincides with the step function outside a narrow interval around the threshold, and rises continuously from 0 to 1 inside it. Using logoids, the task of finding steady states is considerably simplified. A list of regions in phase space comprising all steady states lying close to a threshold is obtained by examining a certain type of matrix called the Logoid-Jacobian. In addition, this matrix leads to the conditions necessary for stability of the steady states. External signals may be conveniently incorporated in the form of Boolean variables. Thus the framework is well suited for studying gene regulatory networks both in single cells and multicellular systems.

Effects of various breeding strategies on diploid drone frequency and quantitative traits in a honey bee population.

When selecting in a finite population of honeybees there is a conflict between gain in a quantitative trait and increasing homozygosity, and therefore the frequency of inviable diploid drones. The consequences when using different mating, import, and selection strategies on diploid drone frequency and genetic gain, was explored with Monte Carlo computer simulations.Within a closed population breeding structure, mass selection gave the highest genetic gain in the quantitative trait, but also the largest increase in percentage diploid drones and queens with unacceptably-low brood viability. Mass selection combined with truncation selection against queens having more than 15% diploid drones gave a comparable genetic gain and was the best strategy of the ones studied to avoid diploid drones. Within-family selection (one replacement per sib group) gave the least genetic gain, and a frequency of diploid drones comparable to random (no) selection. It was intermediate between mass selection and mass selection combined with viability selection concerning the frequency of diploid drones.Insemination with pooled and homogenized semen originating from all breeder queens (30), as compared to natural mating with 12 randomly-selected drones, had little effect on the genetic gain and on the overall frequency of diploid drones (10 to 15% by generation 20).The effect of opening the closed breeding population for the import of external queens every generation, by exchanging breeder queens of lowest performance with a corresponding number of new queens (5, 10and 15 out of 30), was also investigated. Under mass selection (natural mating as well as artificial insemination) the frequency of diploid drones and the proportion of queens discarded were reduced because of low brood viability. However, artificial insemination was superior to natural mating considering the latter criterion. If the imported queens were at the same genetic level for the quantitative trait under selection as the whole breeding population at that generation, or 10% better, the genetic gain was respectively slightly reduced and approximately maintained. If the imported queens were of inferior quality (equal to the initial population) the import of queens slowed genetic progress considerably.