Bioinformatic analysis and functional predictions of selected regeneration-associated transcripts expressed by zebrafish microglia

Abstract

Background Unlike mammals, zebrafish have a remarkable capacity to regenerate a variety of tissues, including central nervous system tissue. The function of macrophages in tissue regeneration is of great interest, as macrophages respond and participate in the landscape of events that occur following tissue injury in all vertebrate species examined. Understanding macrophage populations in regenerating tissue (such as in zebrafish) may inform strategies that aim to regenerate tissue in humans. We recently published an RNA-seq experiment that identified genes enriched in microglia/macrophages in regenerating zebrafish retinas. Interestingly, a small number of transcripts differentially expressed by retinal microglia/macrophages during retinal regeneration did not have predicted orthologs in human or mouse. We reasoned that at least some of these genes could be functionally important for tissue regeneration, but most of these genes have not been studied experimentally and their functions are largely unknown. To reveal their possible functions, we performed a variety of bioinformatic analyses aimed at identifying the presence of functional protein domains as well as orthologous relationships to other species. Results Our analyses identified putative functional domains in predicted proteins for a number of selected genes. For example, we confidently predict kinase function for one gene, cytokine/chemokine function for another, and carbohydrate enzymatic function for a third. Predicted orthologs were identified for some, but not all, genes in species with described regenerative capacity, and functional domains were consistent with identified orthologs. Comparison to other published gene expression datasets suggest that at least some of these genes could be important in regenerative responses in zebrafish and not necessarily in response to microbial infection. Conclusions This work reveals previously undescribed putative function of several genes implicated in regulating tissue regeneration. This will inform future work to experimentally determine the function of these genes in vivo, and how these genes may be involved in microglia/macrophage roles in tissue regeneration.

A Comparative Analysis of Gene and Protein Expression Throughout a Full 28-Day Retinal Regeneration Time-Course in Adult Zebrafish

“…The presence of 4C4+ microglia at 72 hpl also immediately followed the peak of apoptosis in the outer retina from 24 to 36 hpl as evidenced by TUNEL assay ( Supplementary Figure 1 ). Together, these data support a previous suggestion that quantification of 4C4+ microglia could be used as a proxy for retinal neuron damage ( Craig et al, 2008 ; White et al, 2017 ; Ranski et al, 2018 ; Mitchell et al, 2019 ; Issaka Salia and Mitchell, 2020 ).…”

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“…of apoptosis in the outer retina from 24 to 36 hpl as evidenced by TUNEL assay (Supplementary Figure 1). Together, these data support a previous suggestion that quantification of 4C4+ microglia could be used as a proxy for retinal neuron damage (Craig et al, 2008;White et al, 2017;Ranski et al, 2018;Mitchell et al, 2019;Issaka Salia and Mitchell, 2020).…”

A comparative analysis of gene and protein expression in chronic and acute models of photoreceptor degeneration in adult zebrafish

“…Müller glia/macroglia function together with microglia in a number of developmental, disease, and regeneration contexts ( White et al, 2017 ; Mitchell et al, 2019 ; Issaka Salia and Mitchell, 2020 ; Var and Byrd-Jacobs, 2020 ). In 0 h CLL control retinas, microglia appeared to exhibit two distinct morphologies ( Figure 4A ; Supplementary Figure S6A ).…”

“…The role of microglia in retinal development and regeneration is a growing area of study ( White et al, 2017 ; Mitchell et al, 2019 ; Issaka Salia and Mitchell, 2020 ). In a resting state, microglia typically reside in the plexiform layers of the retina, exhibiting a spindle-shaped, ramified morphology ( Nimmerjahn et al, 2005 ).…”