Comparative functional analysis of ribonuclease 1 homologs: molecular insights into evolving vertebrate physiology

Abstract

Pancreatic-type ribonucleases (ptRNases) comprise a class of highly conserved secretory endoribonucleases in vertebrates. The prototype of this enzyme family is ribonuclease 1 (RNase 1). Understanding the physiological roles of RNase 1 is becoming increasingly important, as engineered forms of the enzyme progress through clinical trials as chemotherapeutic agents for cancer. Here we present an in-depth biochemical characterization of RNase 1 homologs from a broad range of mammals (human, bat, squirrel, horse, cat, mouse, and cow) and non-mammalian species (chicken, lizard, and frog). We discover that the human homolog of RNase 1 has a pH optimum for catalysis, ability to degrade double-stranded RNA, and affinity for cell-surface glycans that are distinctly higher than those of its homologs. These attributes have relevance for human health. Moreover, the functional diversification of the ten RNase 1 homologs illuminates the regulation of extracellular RNA and other aspects of vertebrate evolution.

Phenotype of ribonuclease 1 deficiency in mice

“…We note, too, that eRNA secondary structure is thought to be important for its procoagulant activity (Gansler et al 2012). Human RNase 1, which has higher activity against double-stranded RNA than does either mouse RNase 1 or RNase A (Eller et al 2014;Lomax et al 2017), might be better adapted to regulate endogenous eRNA alone.…”

“…Excess RNase A was added, and maximum fluorescence intensity (I max ) was measured. Recombinant murine RNase 1 was used to measure k cat /K M = 8.6 × 10 6 M -1 sec -1 , which is indistinguishable from the value at pH 6.4 reported previously (Lomax et al 2017). This value was, in turn, used to estimate the concentration of ribonuclease in mouse plasma with Equation 1:…”

“…Previously, we showed that the murine RI•RNase 1 complex is highly similar to its human homolog in its three-dimensional structure (Lomax et al 2014). Further, murine RNase 1 exhibits ribonucleolytic activity similar to that of the human enzyme (Lomax et al 2017). Given that human Rnase1 is expressed ubiquitously (Futami et al 1997), we analyzed 18 mouse tissues for Rnase1 expression by qPCR and found the mRNA to be expressed ubiquitously (Fig.…”

Immune Modulation by Human Secreted RNases at the Extracellular Space

“…hRNase1 is the human homolog of the family reference protein, bovine pancreatic RNaseA. A comparative evolutionary analysis indicates a divergent role for non-ruminant RNase1 family members that are unrelated to digestion ( 18 , 142 ). Expression of human RNase1 was detected in almost all tissues ( 143 ).…”

“…The authors considered the potential contribution to RNA clearance by eosinophil secreted RNases. Later studies also indicated an RNA scavenging role for pancreatic-type RNases ( 18 , 19 ). In addition, the secreted proteins displayed immuno-regulatory properties that suggest they could participate in the transmission and amplification of local danger signals ( 17 , 20 ).…”