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Mice enter the post-genome era

The enormous advance in the speed and availability of sequencing technology during recent years means that today we possess data on the genomes of a truly diverse range of organisms. However, we know far less about the related sequences of amino acids and actual proteins. This is the specific area of research of the laboratory of Prof. Pavel Stopka of the Zoology Department who today works at BIOCEV. He recently published a study in the journal Scientific Reports on the proteome of mouse cavities that focused primarily on proteins associated with signalling and immune response.

 

IMG_9372_Snapseed.jpg The team of Prof. Pavel Stopka, Ph.D. (centre) at BIOCEV. Photo: archive of Pavel Stopka.  

 

 

Mice, specifically the laboratory mice that are well known to practically everyone who has ever conducted research in the fields of biology and medicine, represent a very specific subject of study. From a zoological perspective, they are a hybrid of three subspecies of the wild house mouse (the third got into the gene pool more or less accidentally). Other modifications of genetic information in laboratory mice have occurred through inbreeding.

In our research we use the Eastern European house mouse (Mus musculus musculus). Our study is the first in the area of functional mouse genomics that works with wild animals. Most biomedical research is conducted on laboratory mice. But because of crossbreeding and inbreeding these mice have lost many of the characteristics we find in wild mice. It is important for us to discover proteins that are practically unknown in laboratory mice,” says Prof. Pavel Stopka about just one of the unique findings of his research team.

Today, the “mouse” genome is already well mapped. It consists of between 20 – 22,000 sequences that are presumed to encode functional proteins. However, no functional protein has yet been found for the overwhelming majority of these. The current proteome research (of the oral cavity) is a continuation of the long-term interest of Pavel Stopka and his team in the proteins involved in chemical communication, especially intra-species communication. These proteins in mice are found in numerous body fluids and are released on the body’s surface or in urine, tears or saliva. The primary goal of the scientific work is to create a database of the salivary proteome. “For us to use the mouse as a model, we need to know the composition of its proteome. Only then we will be able to focus on individual families and analyse the function of individual proteins” explains Pavel Stopka.  

 

IMG_7319 uprava.jpg The oral cavities of mice frequently come into contact during social interactions. This results in the transfer of chemical signals and often various pathogens as well. Photo: Romana Stopková

 

The work builds upon the long-term interest of Pavel Stopka and his research team in proteins that are associated with chemical communication in mice, especially lipocalins. These proteins serve primarily for intra-species communication and are therefore present in urine, tears and saliva. “During our work we gradually encountered the problem of how to compare the amount of proteins in a sample. We therefore began using liquid chromatography combined with mass spectrometry (LC-MS/MS). The results even surprised us. After thoroughly filtering the data statistically, we identified over 600 different types of protein in saliva, which was more than twice the amount we expected. An even greater surprise was that nearly 21% were gender-specific, occurring much more frequently in males or in females,” says Prof. Stopka of one of the more interesting findings of his research.

And what are some other new discoveries divulged to the scientific community in the article in Scientific Reports? First it was shown that saliva contains a number of proteins with very diverse functions. Some of these function, for example, as transporters of waste products from the nasal epithelium. This function is often performed by lipocalins, which otherwise bind chemical signals (Odorant-binding proteins, OBP).  Toxic-waste  products, which are further transported to the digestive system, are bound to specific receptors called beta-baskets. A number of proteins expressed in the oral cavity also have an immunity function. “We have significantly expanded the spectrum of antimicrobial proteins, especially those from the family of BPI proteins, which disrupt bacterial membranes. This is a very old evolutionary mechanism that protects organisms from pathogens. We are presently testing these proteins in vitro in various cultures including non-pathogenic bacteria,” says Pavel Stopka about the direction of future research.

 

IMG_7300 uprava.jpg Self-grooming is behaviour where mice lick their front paws and then deposit antimicrobial proteins in their fur.  Photo: Romana Stopková

 

As a number of proteins present in mouse salivary gland products take part in immune responses, they can be used as biomarkers for various immune processes in the body. Healthy and sick mice can then easily be identified based only on their different proteomic profiles. Numerous proteins (including certain lipocalins) can be used as biomarkers for the oestrous cycle of a female. “This is quite significant for other biomedical research. Until now, the oestrous cycle has primarily been determined through vaginal flushing. During this procedure however, infections were easily passed into the genital tract, which distorted subsequent findings. By using salivary proteins we can determine the oestrous cycle non-invasively,” explains Stopka. 

Knowledge gained from research on mice may in the future also be helpful for human medical research. Potential biomedical applications are based on the fact that humans (primates) and mice (rodents) share  many plesiomorphies and apomorphies that are typical for placental mammals from the group Euarchontoglires. The similarity of coding sequences between rodents and humans is typically very high, between 95 - 99%. “However, it is often the case that the products of the same gene acquire a different meaning and function in the context of the body of a new group of organisms. Creating a proteome database is an essential first step in the search for all other functional proteins,” concludes Prof. Stopka.

 

Michal Andrle

 

Published: Oct 18, 2016 02:50 PM

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