Spinocerebellar ataxia type 2 (SCA2) is caused by the expansion of a polyglutamine (polyQ) repeat in ataxin-2, the SCA2 gene product. In contrast to other polyQ diseases, intranuclear inclusions are not prominent in SCA2. In animal models with expression of mutant ataxin-2 targeted to Purkinje cells, neuronal dysfunction and morphologic changes are observed without the formation of intranuclear aggregates. In this report, we investigated the mechanisms underlying SCA2 pathogenesis using cellular models. We confirmed that the SCA2 gene product, ataxin-2, was predominantly located in the Golgi apparatus. Deletion of ER-exit and trans-Golgi signals in ataxin-2 resulted in an altered subcellular distribution. Expression of full-length ataxin-2 with an expanded repeat disrupted the normal morphology of the Golgi complex and colocalization with Golgi markers was lost. Intranuclear inclusions were only seen when the polyQ repeat was expanded to 104 glutamines, and even then were only observed in a small minority of cells. Expression of ataxin-2 with expanded repeats in PC12 and COS1 cells increased cell death compared with normal ataxin-2 and elevated the levels of activated caspase-3 and TUNEL-positive cells. These results suggest a link between cell death mediated by mutant ataxin-2 and the stability of the Golgi complex. The formation of intranuclear aggregates is not necessary for in vitro cell death caused by expression of full-length mutant ataxin-2.

Inactivating mutations of the gene encoding parkin are responsible for some forms of autosomal recessive juvenile Parkinson disease. Parkin is a ubiquitin ligase that ubiquitinates misfolded proteins targeted for the proteasome-dependent protein degradation pathway. Using the yeast two-hybrid system and co-immunoprecipitation methods, we identified synaptotagmin XI as a protein that interacts with parkin. Parkin binds to the C2A and C2B domains of synaptotagmin XI resulting in the polyubiquitination of synaptotagmin XI. Truncated and missense mutated parkins reduce parkin-sytXI binding affinity and ubiquitination. Parkin-mediated ubiquitination also enhances the turnover of sytXI. In sporadic PD brain sections, sytXI was found in the core of the Lewy bodies. Since synaptotagmin XI is a member of the synaptotagmin family that is well characterized in their importance for vesicle formation and docking, the interaction with this protein suggests a role for parkin in the regulation of the synaptic vesicle pool and in vesicle release. Loss of parkin could thus affect multiple proteins controlling vesicle pools, docking and release and explain the deficits in dopaminergic function seen in patients with parkin mutations.

Expansion of the polyQ repeat in ataxin-2 results in degeneration of Purkinje neurons and other neuronal groups including the substantia nigra in patients with spinocerebellar ataxia type 2 (SCA2). In animal and cell models, overexpression of mutant ataxin-2 induces cell dysfunction and death, but little is known about steady-state levels of normal and mutant ataxin-2 and cellular mechanisms regulating their abundance. Based on preliminary findings that ataxin-2 interacted with parkin, an E3 ubiquitin ligase mutated in an autosomal recessive form of Parkinsonism, we sought to determine whether parkin played a role in regulating the steady-state levels of ataxin-2. Parkin interacted with the N-terminal half of normal and mutant ataxin-2, and ubiquitinated the full-length form of both wild-type and mutant ataxin-2. Parkin also regulated the steady-state levels of endogenous ataxin-2 in PC12 cells with regulatable parkin expression. Parkin reduced abnormalities in Golgi morphology induced by mutant ataxin-2 and decreased ataxin-2 induced cytotoxicity. In brains of SCA2 patients, parkin labeled cytoplasmic ataxin-2 aggregates in Purkinje neurons. These studies suggest a role for parkin in regulating the intracellular levels of both wild-type and mutant ataxin-2, and in rescuing cells from ataxin-2-induced cytotoxicity. The role of parkin variants in modifying the SCA2 phenotype and its use as a therapeutic target should be further investigated.

Cell migration is essential for regulating many biological processes in physiological or pathological conditions, including embryonic development and cancer invasion. In vitro and in silico studies suggest that collective cell migration is associated with some biomechanical particularities, such as restructuring of extracellular matrix (ECM), stress and force distribution profiles, and reorganization of cytoskeleton. Therefore, the phenomenon could be understood by an in-depth study of cells' behavior determinants, including but not limited to mechanical cues from the environment and from fellow "travelers". This review article aims to cover the recent development of experimental and computational methods for studying the biomechanics of collective cell migration during cancer progression and invasion. We also summarized the tested hypotheses regarding the mechanism underlying collective cell migration enabled by these methods. Together, the paper enables a broad overview on the methods and tools currently available to unravel the biophysical mechanisms pertinent to cell collective migration, as well as providing perspectives on future development towards eventually deciphering the key mechanisms behind the most lethal feature of cancer. I.

Heterogeneous intercellular coupling plays a significant role in mechanical and electrical signal transmission in the heart. Although many studies have investigated the electrical signal conduction between myocytes and nonmyocytes within the heart muscle tissue, there are not many that have looked into the mechanical counterpart. This study aims to investigate the effect of substrate stiffness and the presence of cardiac myofibroblasts (CMFs) on mechanical force propagation across cardiomyocytes (CMs) and CMFs in healthy and heart-attack-mimicking matrix stiffness conditions. The contractile forces generated by the CMs and their propagation across the CMFs were measured using a bio-nanoindenter integrated with fluorescence microscopy for fast calcium imaging. Our results showed that softer substrates facilitated stronger and further signal transmission. Interestingly, the presence of the CMFs attenuated the signal propagation in a stiffness-dependent manner. Stiffer substrates with CMFs present attenuated the signal $24-32% more compared to soft substrates with CMFs, indicating a synergistic detrimental effect of increased matrix stiffness and increased CMF numbers after myocardial infarction on myocardial function. Furthermore, the beating pattern of the CMF movement at the CM-CMF boundary also depended on the substrate stiffness, thereby influencing the waveform of the propagation of CM-generated contractile forces. We performed computer simulations to further understand the occurrence of different force transmission patterns and showed that cell-matrix focal adhesions assembled at the CM-CMF interfaces, which differs depending on the substrates stiffness, play important roles in determining the efficiency and mechanism of signal transmission. In conclusion, in addition to substrate stiffness, the degree and type of cell-cell and cell-matrix interactions, affected by the substrate stiffness, influence mechanical signal conduction between myocytes and nonmyocytes in the heart muscle tissue.