Bioelectric-calcineurin signaling module regulates allometric growth and size of the zebrafish fin

Abstract

The establishment of relative size of organs and structures is paramount for attaining final form and function of an organism. Importantly, variation in the proportions of structures frequently underlies adaptive change in morphology in evolution and maybe a common mechanism underlying selection. However, the mechanism by which growth is integrated within tissues during development to achieve proper proportionality is poorly understood. We have shown that signaling by potassium channels mediates coordinated size regulation in zebrafish fins. Recently, calcineurin inhibitors were shown to elicit changes in zebrafish fin allometry as well. Here, we identify the potassium channel kcnk5b as a key player in integrating calcineurin’s growth effects, in part through regulation of the cytoplasmic C-terminus of the channel. We propose that the interaction between Kcnk5b and calcineurin acts as a signaling node to regulate allometric growth. Importantly, we find that this regulation is epistatic to inherent mechanisms instructing overall size as inhibition of calcineurin is able to bypass genetic instruction of size as seen in sof and wild-type fins, however, it is not sufficient to re-specify positional memory of size of the fin. These findings integrate classic signaling mediators such as calcineurin with ion channel function in the regulation of size and proportion during growth.

Integrated K+channel and K+-Cl−cotransporter functions regulate fin proportionality in zebrafish

“…Clonal analysis on fin tissue verified that the guide RNAs successfully introduced deletions and/or insertions at the targeted sites (Figure 7d). As would be expected based on our earlier work (Perathoner, et al, 2014), (Daane, et al, 2018), targeted inactivation of kcnk5b had no effect in a wild-type genetic background (Figure 7a). However, CRISPR-mediated inactivation of kcnk5b in a heterozygous slc12a7a/schleier background produced fish with reduced fin and barbel length (Figure 7c).…”

“…This finding resembles the phenotype of the another longfin (alf) or pfau long-finned mutants, which exhibit comparable changes in fin skeletal patterning and are caused by gain of function-alterations in Kcnk5b (Perathoner, et al, 2014). Given the phenotypic similarity between these mutants and that our previous work linked alf/kcnk5b function to bioelectric signaling in development (Daane, et al, 2018), we chose to further characterize schleier as it might reveal mechanistic aspects of bioelectric signaling in size regulation.…”

“…We have previously demonstrated that activating mutations in the potassium channel kcnk5b caused patterned overgrowth of fins and barbels, and that absence of kcnk5b had no effect on fin phenotype (Perathoner, et al, 2014), (Daane, et al, 2018). Given the similarity between the kcc4a and kcnk5b overgrowth phenotypes, we hypothesized that they might work in a similar pathway or act in parallel to regulate growth.…”

Recent advancements in understanding fin regeneration in zebrafish

“…This intriguingly indicates that isometric fin growth is controlled by bioelectrical signals. Segment length and fin size is also increased when the protein phosphatase calcineurin is inhibited (Daane et al, 2018;Kujawski et al, 2014). Surprisingly, calcineurin inhibition has no effect on growth in knck5b (alf ) mutants, and calcineurin was suggested to directly modify conductance of the channel (Daane et al, 2018).…”

“…Segment length and fin size is also increased when the protein phosphatase calcineurin is inhibited (Daane et al, 2018;Kujawski et al, 2014). Surprisingly, calcineurin inhibition has no effect on growth in knck5b (alf ) mutants, and calcineurin was suggested to directly modify conductance of the channel (Daane et al, 2018). How bioelectrial signals are induced and regulated and how knck5b/calcineurin signals regulate fin growth, will be interesting avenues for future research.…”