Browsing by Author "Holsinger, RMD"

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    Intrinsic microtubule GTP-cap dynamics in semi-confined systems: kinetochore–microtubule interface
    (Springer Nature, 2012-10-18) Buljan, VA; Holsinger, RMD; Hambly, BD; Banati, RB; Ivanova, EP
    In order to quantify the intrinsic dynamics associated with the tip of a GTP-cap under semi-confined conditions, such as those within a neuronal cone and at a kinetochore–microtubule interface, we propose a novel quantitative concept of critical nano local GTP-tubulin concentration (CNLC). A simulation of a rate constant of GTP-tubulin hydrolysis, under varying conditions based on this concept, generates results in the range of 0-420 s−1. These results are in agreement with published experimental data, validating our model. The major outcome of this model is the prediction of 11 random and distinct outbursts of GTP hydrolysis per single layer of a GTP-cap. GTP hydrolysis is accompanied by an energy release and the formation of discrete expanding zones, built by less-stable, skewed GDP-tubulin subunits. We suggest that the front of these expanding zones within the walls of the microtubule represent soliton-like movements of local deformation triggered by energy released from an outburst of hydrolysis. We propose that these solitons might be helpful in addressing a long-standing question relating to the mechanism underlying how GTP-tubulin hydrolysis controls dynamic instability. This result strongly supports the prediction that large conformational movements in tubulin subunits, termed dynamic transitions, occur as a result of the conversion of chemical energy that is triggered by GTP hydrolysis (Satarić et al., Electromagn Biol Med 24:255–264, 2005). Although simple, the concept of CNLC enables the formulation of a rationale to explain the intrinsic nature of the “push-and-pull” mechanism associated with a kinetochore–microtubule complex. In addition, the capacity of the microtubule wall to produce and mediate localized spatio-temporal excitations, i.e., soliton-like bursts of energy coupled with an abundance of microtubules in dendritic spines supports the hypothesis that microtubule dynamics may underlie neural information processing including neurocomputation. © 2012, Springer Nature
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    Intrinsic synergistic-topological mechanism versus synergistic-topological matrix in microtubule self-organization
    (BioMed Central Ltd, 2014-12-04) Buljan, VA; Holsinger, RMD; Hambly, BD; Kanellis, VG; Matar, E; Larkin, X; Liu, GJ; Bohorquez-Florez, JJ; Banati, RB
    Background In this body of work we investigate the synergistic-topological relationship during self-organization of the microtubule fiber in vitro, which is composed of straight, axially shifted and non-shifted, acentrosomal microtubules under crowded conditions. Methods We used electron microscopy to observe morphological details of ordered straight microtubules. This included the observation of the differences in length distribution between microtubules in ordered and non-ordered phases followed by the observation of the formation of interface gaps between axially shifted and ordered microtubules. We performed calculations to confirm that the principle of summation of pairwise electrostatic forces act between neighboring microtubules all their entire length. Results We have shown that the self-organization of a microtubule fiber imposes a variety of topological restrictions onto its constituting components: (a) tips of axially shifted neighboring microtubules are not in direct contact but rather create an ‘interface gap’; (b) fibers are always composed of a restricted number of microtubules at given solution conditions; (c) the average length of microtubules that constitute a fiber is always shorter than that of microtubules outside a fiber; (d) the length distribution of microtubules that constitute a fiber is narrower than that of microtubules outside a fiber and this effect is more pronounced at higher GTP-tubulin concentrations; (e) a cooperative motion of fiber microtubules due to actualization of the summation principle of pairwise electrostatic forces; (f) appearance of local GTP-tubulin depletion immediately in front of the tips of fiber microtubules. Conclusion Overall our data indicate that under crowded conditions in vitro, the self-organization of a microtubule fiber is governed by an intrinsic synergistic-topological mechanism, which in conjunction with the topological changes, GTP-tubulin depletion, and cooperative motion of fiber constituting microtubules, may generate and maintain a ‘synergistic-topological matrix’. Failure of the mechanism to form biologically feasible microtubule synergistic-topological matrix may, per se, precondition tumorigenesis. © 2014 Authors
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    Selective, high-contrast detection of syngeneic glioblastoma in vivo
    (Springer Nature, 2020-06-19) Banati, RB; Wilcox, P; Xu, R; Yin, G; Si, E; Son, ET; Shimizu, M; Holsinger, RMD; Parmar, A; Zahra, D; Arthur, A; Middleton, RJ; Liu, GJ; Charil, A; Graeber, MB
    Glioblastoma is a highly malignant, largely therapy-resistant brain tumour. Deep infiltration of brain tissue by neoplastic cells represents the key problem of diffuse glioma. Much current research focuses on the molecular makeup of the visible tumour mass rather than the cellular interactions in the surrounding brain tissue infiltrated by the invasive glioma cells that cause the tumour’s ultimately lethal outcome. Diagnostic neuroimaging that enables the direct in vivo observation of the tumour infiltration zone and the local host tissue responses at a preclinical stage are important for the development of more effective glioma treatments. Here, we report an animal model that allows high-contrast imaging of wild-type glioma cells by positron emission tomography (PET) using [18 F]PBR111, a selective radioligand for the mitochondrial 18 kDa Translocator Protein (TSPO), in the Tspo−/− mouse strain (C57BL/6-Tspotm1GuMu(GuwiyangWurra)). The high selectivity of [18 F]PBR111 for the TSPO combined with the exclusive expression of TSPO in glioma cells infiltrating into null-background host tissue free of any TSPO expression, makes it possible, for the first time, to unequivocally and with uniquely high biological contrast identify peri-tumoral glioma cell invasion at preclinical stages in vivo. Comparison of the in vivo imaging signal from wild-type glioma cells in a null background with the signal in a wild-type host tissue, where the tumour induces the expected TSPO expression in the host’s glial cells, illustrates the substantial extent of the peritumoral host response to the growing tumour. The syngeneic tumour (TSPO+/+) in null background (TSPO−/−) model is thus well suited to study the interaction of the tumour front with the peri-tumoral tissue, and the experimental evaluation of new therapeutic approaches targeting the invasive behaviour of glioblastoma. © 2020, The Author(s).
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    Up-regulation of matrix metallopeptidase 12 in motor neurons undergoing synaptic stripping
    (Elsevier B.V., 2014-08-22) Sajjan, S; Holsinger, RMD; Fok, S; Ebrahimkhani, S; Rollo, JL; Banati, RB; Graeber, MB
    Axotomy of the rodent facial nerve represents a well-established model of synaptic plasticity. Post-traumatic “synaptic stripping” was originally discovered in this system. We report upregulation of matrix metalloproteinase MMP12 in regenerating motor neurons of the mouse and rat facial nucleus. Matrix metalloproteinases (matrix metallopeptidases, MMPs) are zinc-binding proteases capable of degrading components of the extracellular matrix and of regulating extracellular signaling networks including within synapses. MMP12 protein expression in facial motor neurons was enhanced following axotomy and peaked at day 3 after the operation. The peak of neuronal MMP12 expression preceded the peak of experimentally induced synaptic plasticity. At the same time, MMP12 redistributed intracellularly and became predominantly localized beneath the neuronal somatic cytoplasmic membrane. Both findings point to a role of MMP12 in the neuronal initiation of the synaptic stripping process. MMP12 is the first candidate molecule for such a trigger function and has potential as a therapeutic target. Moreover, since statins have been shown to increase the expression of MMP12, interference with synaptic stability may represent one mechanism by which these widely used drugs exert their side effects on higher CNS functions. © 2014 by Elsevier Ltd.