miRNAs in ancient tissue specimens of the Tyrolean Iceman

The analysis of nucleic acids in ancient samples is largely limited to DNA. Small noncoding RNAs (microRNAs) are known
to be evolutionary conserved and stable. To gain knowledge on miRNAs measured from ancient samples, we profiled
microRNAs in cryoconserved mummies. First, we established the approach on a World War One warrior, the
“Kaiserj€ager”, which has been preserved for almost one century. Then, we profiled seven ancient tissue specimens
including skeletal muscle, stomach mucosa, stomach content and two corpus organ tissues of the 5,300-year-old copper
age mummy Iceman and compared these profiles to the presence of organ-specific miRNAs in modern tissues. Our
analyses suggest the presence of specific miRNAs in the different Iceman’s tissues. Of 1,066 analyzed human miRNAs, 31
were discovered across all biopsies and 87 miRNAs were detected only in a single sample. To check for potential
microbiological contaminations, all miRNAs detected in Iceman samples and not present in ancient samples were
mapped to 14,582 bacterial and viral genomes. We detected few hits (3.9% of miRNAs compared with 3.6% of
miRNAs). Interestingly, the miRNAs with higher abundance across all ancient tissues were significantly enriched for
Guanine (P value of 10–13) and Cytosine (P value of 10–7). The same pattern was observed for modern tissues.
Comparing miRNAs measured from ancient organs to modern tissue patterns highlighted significant similarities, e.g.,
formiRNAs present in themuscle. Our first comprehensive analysis of microRNAs in ancient human tissues indicates that
these stable molecules can be detected in tissue specimens after 5,300 years.