Genomic conservation of erythropoietic microRNAs (erythromiRs) in white-blooded Antarctic icefish

Abstract

White-blooded Antarctic crocodile icefish are the only vertebrates known to lack functional hemoglobin genes and red blood cells throughout their lives. We do not yet know, however, whether extinction of hemoglobin genes preceded loss of red blood cells or vice versa, nor whether erythropoiesis regulators disappeared along with hemoglobin genes in this erythrocyte-null clade. Several microRNAs, which we here call erythromiRs, are expressed primarily in developing red blood cells in zebrafish, mouse, and humans. Abrogating some erythromiRs, like Mir144 and Mir451a, leads to profound anemia, demonstrating a functional role in erythropoiesis. Here, we tested two not mutually exclusive hypotheses: 1) that the loss of one or more erythromiR genes extinguished the erythropoietic program of icefish and/or led to the loss of globin gene expression through pseudogenization; and 2) that some erythromiR genes were secondarily lost after the loss of functional hemoglobin and red blood cells in icefish. We explored smallRNA transcriptomes generated from the hematopoietic kidney marrow of four Antarctic notothenioids: two red-blooded species (bullhead notothen Notothenia coriiceps and emerald notothen Trematomus bernacchii) and two white-blooded icefish (blackfin icefish Chaenocephalus aceratus and hooknose icefish Chionodraco hamatus). The N. coriiceps genome assembly anchored analyses. Results showed that, like the two red-blooded species, the blackfin icefish genome possessed and the marrow expressed all known erythromiRs. This result indicates that loss of hemoglobin and red blood cells in icefish was not caused by loss of known erythromiR genes. Furthermore, expression of only one erythromiR, mir96, appears to have been lost after the loss of red blood cells and hemoglobin – expression was not detected in the erythropoietic organ of hooknose icefish but was present in blackfin icefish. All other erythromiRs investigated, including mir144 and mir451a, were expressed by all four species and thus are present in the genomes of at least the two white-blooded icefish. Our results rule out the hypothesis that genomic loss of any known erythromiRs extinguished erythropoiesis in icefish, and suggest that after the loss of red blood cells, few erythromiRs experienced secondary loss. Results suggest that functions independent of erythropoiesis maintained erythromiRs, thereby highlighting the evolutionary resilience of miRNA genes in vertebrate genomes.

Embryogenesis and early skeletogenesis in the antarctic bullhead notothen, Notothenia coriiceps

“…For comparison, the sub‐Antarctic Patagonian rockcod Patagonotothen ramsayi , when cultured at its habitat temperature of 5 °–6 °C, hatches in just 24 days (Arkhipkin et al, ). The first recorded natural mating of Antarctic fish in the Northern Hemisphere involved a pair of Chionodraco hamatus (hooknose icefish, see Desvignes et al, for etymology) that had been cultured for three years at ‐1 °C in an Italian aquarium; mating occurred in February and hatching in late June, about 19 weeks after fertilization (Ferrando et al, ). In the Ross Sea, Pleuragramma antarcticum appears to hatch in November, about 4 months after fertilization (Vacchi et al, ).…”

Antarctic blackfin icefish genome reveals adaptations to extreme environments

“…miRNA sequencing and annotation. Organs for miRNA annotation originated from the same male specimen of C. aceratus used in the study of erythropoietic miRNAs in Antarctic icefishes 69 . Samples of pronephric (head) kidney, pectoral girdle bone, heart ventricle, pectoral adductor muscle and skeletal muscle were dissected and stored in RNAlater at −80 °C until further use at the University of Oregon.…”

miRNA analysis with Prost! reveals evolutionary conservation of organ-enriched expression and post-transcriptional modifications in three-spined stickleback and zebrafish

“…Heart also displayed a significant number of evolutionarily-conserved, organ-enriched miRNAs (12 miRNAs) (Figure 3B, D, E). Heart-enriched miRNAs included mature products of the well-described vertebrate cardiac myomiR genes mir1, mir133 , and mir499 55,67,68,74–77 and erythromiRs mir144 and mir451 43 . The former group participate in muscle formation and function, and the latter may reflect the presence of red blood cells in the heart ventricle at the time of RNA extraction.…”

“…Earlier versions of Prost! have been used to annotate zebrafish and spotted gar miRNAs 41,42 , as well as to identify erythromiRs in white-blooded Antarctic icefish 43 .…”