Site-Directed Mutagenesis and Site-Specific Binding Analysis of Calmodulin (CaM)

Calcium signaling is a major regulatory system in cells and a crucial part of cell biology. An important element in the decoding of intracellular calcium concentration into downstream processes is the ubiquitous and highly conserved calcium binding protein calmodulin (CaM) which can bind to and modulate the function of hundreds of different target proteins, regulating such processes as synaptic plasticity, gene expression and electrical signaling. The biophysical characterization of binding affinity and cooperative interactions between each of calmodulin’s four EF-hand calcium binding sites is essential for understanding calcium signaling. Highly conserved amino acid sequence differences in the ion binding loops of the EF-hands give each site unique affinity for calcium. EF-hands are almost always found in pairs, where binding to one of the sites affects the affinity of the paired site. We have used spectroscopy to measure site-specific binding in each of the paired binding sites in the CaM N-lobe, along with site-directed mutagenesis, to study the contributions of individual amino acids to the ion binding affinity in the mutated site (cis effects) and in the neighboring site (trans effects). Of the twelve amino acids in the binding loops, five are different between Site 1 and Site 2. We constructed proteins with substituted individual residues from Site 1 to Site 2. CaM with the full Site 1 sequence in both Site 1 and Site 2 shows significant changes in affinity and binding characteristics in both sites. To investigate the contributions of the individual amino acid differences, we made intermediate mutants containing individual amino acid changes in Site 2. The cis-effects of the intermediate mutations on the mutated site, Site 2, seem to be independent and additive, whereas the trans-effects on the non-mutated Site 1 showed unexpected dependence on combinations of amino acid changes in Site 2.Neuroscienc

Distinguishing the commonsense senses

This paper proposes a methodological strategy to investigate the question of the individuation of the senses both from a commonsensical and a scientific point of view. We start by discussing some traditional and recent criteria for distinguishing the senses and argue that none of them taken in isolation seems to be able to handle both points of views. We then pay close attention to the faculty of hearing which offers promising examples of the strategy we pursue of combining commonsense and science. In particular, we argue in favour of a distinction between two kinds of modes of presentation associated with sound perception: a mechanical mode of presentation that makes sounds perceptible by other modalities than audition such as touch and a tonal mode of presentation that makes people conceive of sounds as the proper objects of auditory perception

L’interprétation ordinaire, entre simulation et méta-représentation

Cet essai porte sur quelques aspects de l’opposition entre un modèle perceptif de la communication, selon lequel le témoignage est une source de connaissance directe du fait témoigné, et un modèle inférentiel de la communication, selon lequel le témoignage implique de la part du récepteur une inférence à partir des signes utilisés, des états mentaux de l’émetteur et du reste du contexte. À partir d’une réflexion sur la nature de la capacité de méta-représentation, et sur sa dépendance à l’égard des capacités de perception sociale et de simulation, je montre que l’attribution d’états mentaux souvent complexes à l’émetteur n’empêche pas le fonctionnement d’un mode primitif de communication dans lequel le récepteur hérite directement de l’information transmise par l’émetteur. La compréhension, aussi réflexive soit-elle, reste enracinée dans la tendance naturelle à accepter ce que l’émetteur cherche à nous faire savoir.In this essay, I examine some aspects of the debate between a perceptual model of communication, according to which testimony is a source of knowledge about the communicated fact, and an inferential model of communication, according to which testimony requires from the hearer an inference from the used signs, the speaker’s mental states, and other features of the context. From a reflection on the nature of the capacity for metarepresentation, and its dependence on the capacities of social perception and simulation, I show that the ascription of even complex mental states to the speaker does not preclude communication from functioning on a mode in which the hearer directly inherits the information transmitted by the speaker. Understanding is grounded on the natural tendency to accept what is intelligibly presented to us as true

Cross-Border Cooperation Programme As an Impetus for Establishment of New Modes of Governance in Croatia

Generated as a response to the requirements in managing European cross border cooperation programmes, Neighbourhood Programme Slovenia-Hungary-Croatia 2004-2006 (NP) in particular served as an impetus for establishment of new modes of governance in Croatia. Together with Hungary and Slovenia, Croatia participates in this trilateral Programme as an external EU border candidate state. The NP is implemented as a grant scheme in Croatia for the first time requiring many institutional changes for its implementation. On all levels of government, the NP contributed to the establishment of stronger relations among institutions, horizontally and vertically, and a transfer of know-how in preparation and implementation from experienced partners, Slovenia and Hungary. Such a complex programme generated also problems due to weak administrative capacity, different sources of financing and socio-economic discrepancies. On national level, weak capacity of the Croatian Ministry of Foreign Affairs and European Integration (MFAEI) has directly influenced the initiation and implementation of NP and difficulties in the decision-making process occurred. The relationship between MFAEI and the Delegation of the European Commission was unclear and there were difficulties in developing new administrative procedures. Additionally, insufficiently transparent information dissemination mechanisms through different government levels influenced the quality and selection of project proposals in certain counties. Socio-cultural differences in Croatia were insufficiently considered while setting up the coordination system resulting in the extent and quality of submitted proposals and their success in the selection procedure. Some regions showed successful coordination on county level contributing to selection of greater number of proposals from those counties, while in others, there was a lack of capacity in preparing coherent project proposals. In accordance with the presented first Croatian experiences in the trilateral European cross-border programme, the following can be concluded: In terms of socio-developmental impacts, the joint-project proposal preparation process of the cross border partners created intensive inter-institutional cooperation and inter-institutional communication. However, only successful project proposals result in inter-institutional collaboration through joint implementation of projects, which is considered fundamental to the creation of a network society and building of social capital.

The Dynamics of Deictic Thoughts

Defense of a non-psychological dynamics of demonstrative thought

Qui a peur des qualia corporels?

Qualia, conceived as intrinsic properties of experiences, are not always welcomed by materialists, who prefer to see them as intentional properties presented in our experience. I ask whether this form of reductionism applies to the qualia of bodily awareness. According to the standard materialist theory, the intentional object of pain experience, for instance, is a bodily damage. This theory, though, is unable to account for the phenomenal difference between feeling pain "inside" and perceiving it "outside" (seeing oneself or another in pain). I sketch another reductionist analysis which is compatible with materialism, and according to which the intentional object of bodily awareness, unlike that of external perception, constitutively depends on the subject's experience

Is Memory Purely Preservative?

Dans cet essai, je me demande si la mémoire est une source de connaissance authentique, ou au contraire si elle doit être considérée comme une manière de maintenir en place une connaissance acquise par d'autres moyens épistémiques. Je me prononce en faveur de la première option