Yeast Models Of Parkinson Disease Biology Essay

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Parkinson's disease (PD) is the most common neurodegenerative movement disorder, characterized by a classical triad of motor symptoms [1]. The main focus of this disease is on the progressive degeneration of the dopaminergic neurons in the substantia nigra pars compacta. Although this pathology may explain the motor symptoms it is not thought to underlie other, non-motor, symptoms associated with PD. The pathology is not restricted to the substantia nigra, also other regions of the brain are affected [2].

The neurons in affected regions exhibit abnormal cytoplasmic protein inclusions that are named Lewy bodies (LB) and Lewy neuritis (LN), which are protein aggregates localized in the cell body or neuronal processes, respectively [2,3]. A major constituent of these aggregates is α-synuclein, a natively unfolded protein known to be present in nerve terminals and it has been suggested to play a role in vesicular transport [4,5]. The α-synuclein in LBs and LNs is misfolded and evidence points to this protein as a key player in PD pathogenesis and other neurodegenerative disorders, collectively termed as synucleinopathies [3,4,5].

Missense mutations in the α-synuclein gene are associated with autosomal dominant PD [6]. However, the mechanisms through which mutations lead to PD disease are unknown [7]. Multiplications of the α-synuclein gene are also linked to PD and similar, in several animal models of PD, increased levels of α-synuclein lead to neurodegeneration. This suggests that a simple increase in protein levels might be enough to cause PD pathology. Although the exact link between α-synuclein inclusions and cytotoxicity is unclear, it has been suggested that nonfibrillar dimers and oligomers of α-synuclein, intermediates for LB formation, play an important role in neurodegeneration. Therefore, elucidating mechanisms underlying the cytotoxic effects of α-synuclein may be essential for the development of therapies that can ameliorate of prevent synucleinopathies [4,7].

Summary of seminar

Inside the cell it is very crowded because of the very high concentration of proteins. A substantional fraction of newly synthesized proteins misfold, but the cell has developed quality control mechanisms that can refold or degradate these proteins. When this does not happen, the misfolded proteins can form aggregates which can cause a loss of function or gain of toxic function in the central nervous system. The question is: why, how and when do neurons die? And how can we prevent neurons from dying prematurely?

In the population 0.2% has Parkinson's disease (PD) and in the older population (over 65 years of age) this percentage rises till 1-2%. Age is a major risk factor of this disease. The most important symptoms of PD are a resting tremor, rigidity and difficulties with initiating movements. These symptoms are caused by the loss of dopaminergic neurons. In 1997 the first gene, a-synuclein, associated with PD was identified. Since then there are 16 more genes identified that play a role in PD. However, a-synuclein is one of the major players in the disease, because the Lewy bodies, which are seen in the brains of PD patients, consist of this small protein. A-synuclein is a presynaptic protein abundantly expressed in the brain. However, the function of this long-lived protein is unknown. A fresh view of Parkinsonism revealed that this disease is not limited to the motor system, but affects several brain areas. Do the Lewy bodies cause the neuronal cell loss in these brain areas? At present, they do not think that Lewy bodies are causing (the most important) neuronal cell loss.

To study this, a yeast model was used to investigate the a-synuclein protein. This yeast model was chosen because of the simplicity of this organism. It has a relatively small genome and 60% of the yeast genes have robust homologues with the human genome. The most important reason is the availability of powerful genetic tools. The yeast model may help to understand the basic biology and thereafter it may be translated to other models. It was already shown that individuals with triplications and duplications of the a-synuclein gene develop PD. Different genetic screens were done to study a-synulcein in the yeast model. A synthetic lethal screen was done and it revealed genes that enhance the toxocity of a-synuclein. With an overexpression screen genes were identified that can suppress or enhance the toxicity of a-synuclein. A small molecule screen revealed a class of compounds that reduces the a-synuclein toxicity. These compounds restore the membrane localization and the Endoplasmatic Reticulum (ER)-to-Golgi trafficking of a-synuclein. Remarkably, they also rescue the a-synuclein-induced neurodegeneration in worms and primary neuronal cultures. Thus, a simple organism can teach us lot about the basic molecular mechanisms in PD.

Monomers of a-synuclein form dimers, which form oligomers, which form the final aggregates, the Lewy bodies. If Lewy bodies are not the most important contributors to the neuronal cell loss, which is /are the cytotoxic species? This question was studied using mammalian cells. The dimers and oligomers are not visible after immune histochemistry, so a technique called biomolecular fluorescence complementation was used. Thus, they showed the formation of dimers and oligomers. They became visible, because the a-synuclein protein was tagged with split GFP and when two proteins interacted the GFP fluoresced. The next step was to investigate which genes effect this interaction. An RNA interference screen was done and it revealed different kinases and phosphatases that either increased or decreased the fluorescence of GFP. Consequently, there are highly conserved different pathways that affect the way that the cells maintain protein homeostasis, or proteostasis.

Conclusion /discussion

Although the etiologies of neurodegenerative disorders may be diverse, a common feature is the occurrence of proteinaceous deposits. A crucial role of protein misfolding in the pathogenic process is established by different approaches coming from genetic, animal modeling and biophysics [6]. The identification of genes responsible for familial variants of neurodegenerative diseases, such as α-synuclein and huntingtin (Huntington Disease, HD), is a major step towards understanding the underlying mechanism. The α-synuclein and huntingtin proteins are major components of the cytoplasmic and nuclear inclusions observed in, respectively, PD and HD [6, 8]. However, genome wide screens performed in yeast revealed that genes enhancing the toxicity of α-synuclein did not overlap with the identified genes enhancing huntingtin toxicity, suggesting a distinct underlying, pathogenic mechanism [8].

One may argue that yeast as a model for studying neurodegeneration has its obvious limitations, starting simply with the argument that some genes involved in modulating neurodegeneration may not be present in the yeast genome. The observed toxicity in yeast may also be inherently different from cellular toxicity in neurodegeneration [9]. However, a study in a yeast strain expressing higher levels of α-synuclein identified single compounds that could antagonize α-synuclein-mediated toxicity in yeast, nematode neurons and primary rat neuronal cultures. This suggests that they are acting on deeply rooted biological processes that have been conserved for a billion years of evolution. Thus, this may strengthen the use of the simple yeast model to elucidate complex biological processes in human diseases [10].

Other research using mammalian cells focused on the idea that the oligomeric and dimeric α-synuclein intermediates are more toxic than the inclusions themselves [7, 11]. It has been suggested that cellular processes that lead to increased formation of dimers/oligomers or decreased clearance of these intermediate species are associated with α-synuclein-mediated cytotoxicity [7]. By using the bimolecular fluorescence complementation technique the formation of dimers and oligomers could be directly visualized, but this technique does not directly identify novel interacting partners [11]. Therefore, the first priority of genetic screens is the identification of modifiers, capable of reducing the dimerization and/or oligomerization [12].

In synucleinpathies heterogeneity of symptoms and disease progression is observed, which poses large problems for the discovery of novel targets for treatment [12]. How can the knowledge from current studies be translated into development of therapeutic strategies? The compound screens in yeast successfully indentified inhibitors of α-synuclein toxicity, which may be therapeutic applicable in the future [12]. Investigating the precise nature of α-synuclein toxic species opens new avenues for using these species as a thereapeutic target for synucleinopathies [7].

Personal highlights of the seminar

When I first saw the title of this seminar I thought: 'What does yeast have to do with Parkinson disease?' Before I visited the seminar I did not realize that other, simpler model organisms could learn us that much about complex biological processes. I was fascinated by this and that is also the reason I emphasized the great contribution the yeast model has in the search of novel targets to intervene in synucleinopathies. However, the validity of using the yeast model will only be established by testing of the targets in more physiologically relevant models, such as rats and mice. Although the yeast model may have a great contribution, I do not think that it could reduce the use of animal models.