# Quantitative Immunofluorescence Mapping of HSP70's Neuroprotective Effects in FUS-ALS Mouse Models
## 1 Introduction  
Amyotrophic lateral sclerosis (ALS) is one of the most common neurodegenerative diseases, develops rapidly and results in fatal outcomes [1]. Despite its clinical symptoms and important regulatory points in skeletal muscles, ALS is primarily neuronal damage [2]. A large number of both familial and, to a lesser extent, sporadic ALS cases refer to mutations of the fused in sarcoma/translocated in liposarcoma gene (FUS). For instance, FUS-associated disease is second most common among ALS cases in China [3] and fourth in Western Europe [4], although there are certain regions where FUS mutations in ALS are seldom found [5, 6].  
Molecular chaperones maintain protein homeostasis in cells, thereby taking part in any reaction to neuronal damage, including that in ALS [7]. In FUS-associated ALS, the damage involves aggregation of mutant FUS gene product, aberrant FUS RNA binding protein, and is also managed by molecular chaperones [8]. Among the diverse group of molecular chaperones, heat shock 70 kDa proteins (HSP70) are distinguished for their crucial role of protein quality control in cell populations of nervous tissue [9, 10]. HSP70 have already been shown to prevent aberrant FUS aggregation in affected neurons [8]. Therefore, employing HSP70 upregulation for preventing tissue pathology development in aberrant FUS presence is an aim of high relevance.  
Plausible mechanisms of HSP70-FUS interaction are not comprehensively elucidated yet. Pathogenic FUS leaves its normal nuclear localization for cytoplasm [11]. There, aberrant FUS joins to stress granules [12], i.e., cytoplasmic condensates in the cytoplasm, blocking their functioning [13] or turning into solid state from liquid droplets of FUS, as shown in vitro [14]. Normally, HSP70 should prevent FUS from this transition due to its phase separation potential [15]. However, it is not clear why this potential of normal HSP70 cannot protect neurons in case of FUS-associated ALS development.  
Such a failure of molecular chaperoning can result from a generally low activity of HSP70 in ALS because natural heat shock factor (HSF1) mediated HSP70 upregulation is compromised [16]. Here, we decided to address if the enhancement of the HSP70 expression may be beneficial in FUS-associated pathology. In this regard, we used a murine model with human _FUS [11-359]_ gene insertion [17,18] co-expressed with human HSP70.  
Nevertheless, HSP70 overexpression can be equivocal as different HSP70 isoforms are situated in various cellular locations and act differently in ALS in particular. In cytosol, Hsp70 member 1A (HSPA1A) is considered as the key isoform, and its role for protein quality control was already reported [10]. Further, [PERSON] and colleagues showed HSPA1A to be efficient against development of SOD1-associated ALS [19]. We designed this study to regard cellular changes in nervous tissue due to _FUS [11-359]_-HSPA1A_ interaction in vivo when both genes are expressed together.  
Current research belongs to a series of studies in FUS-related ALS pathogenetic mechanisms in various structures of the central nervous system.