PRSS56 activity is required during both the vision-independent and dependent stages of ocular growth.

<p>To determine the temporal window critical for the PRSS56-mediated effect on ocular axial growth, <i>Prss56</i> conditional mutant mice (<i>Prss56</i>^<i>F/F</i>) were crossed to mice expressing the ubiquitous inducible Ubc-Cre recombinase (<i>Ubc-Cre</i>^<i>ERT2</i>). (<b>A</b>) Schematic of tamoxifen treatment at distinct developmental stages preceding and following the opening of the eyes. Tamoxifen injection at two different time points, P6 and P8, was performed to ablate <i>Prss56</i> after the earliest detectable effect of mutant <i>Prss56</i> on ocular axial length. (<b>B</b>-<b>D</b>) OCT-based ocular biometry demonstrates that following tamoxifen injection, <i>Prss56</i>^<i>F/F</i><i>; Ubc-Cre</i>^<i>ERT2</i> mice display a significantly reduced ocular axial length (<b>B</b>) and increased retinal thickness (<b>C</b>) and a significant decrease in VCD (<b>D</b>) compared to the control <i>Prss56</i>^<i>F/+</i>; Ubc-Cre^<i>ERT2</i> mice (measured at P17). The ocular axial length, retinal thickness, and VCD of uninjected <i>Prss56</i>^<i>F/F</i><i>; Ubc-Cre</i>^<i>ERT2</i> <i>and Prss56</i>^<i>F/+</i><i>; Ubc-Cre</i>^<i>ERT2</i> mice were indistinguishable. Administration of tamoxifen at P6 caused a greater decrease in ocular axial length compared to administration at P8 suggesting a requirement for continuous PRSS56 activity during ocular development to sustain normal ocular growth, N = 6 to 8 per group for <b>A</b> and <b>B</b>. (<b>E</b>-<b>G</b>) OCT measurements demonstrate that following tamoxifen injection at P13 (a time point when the eyes are open), <i>Prss56</i>^<i>F/F</i><i>; Ubc-Cre</i>^<i>ERT2</i> mice display a slight but significant decrease in ocular axial length (<b>E</b>) and increase in retinal thickness (<b>F</b>) at P30 and P45. Reduced ocular axial length was associated with a significant decrease in VCD in <i>Prss56</i>^<i>F/F</i><i>; Ubc-Cre</i>^<i>ERT2</i> mice compared to <i>Prss56</i>^<i>F/+</i><i>; Ubc-Cre</i>^<i>ERT2</i> and uninjected controls at P30 and P45, N = 5 to 10 per group (<b>G</b>). <b>(H</b>-<b>J)</b> Ocular biometry following tamoxifen injection at the beginning of a critical emmetropization period (P18) shows that the ocular axial length is not significantly different between the <i>Prss56</i>^<i>F/F</i><i>; Ubc-Cre</i>^<i>ERT2</i> and control mice at any of the three ages examined (P30, P45, and P60). However, <i>Prss56</i>^<i>F/F</i><i>; Ubc-Cre</i>^<i>ERT2</i> mice display a slightly thicker retina and significantly reduced VCD compared to the control groups. (<b>K</b>) Following <i>Prss56</i> ablation at P13 and P18, the eyes display a decrease in the combined value of retinal thickness and VCD. (<b>L</b>) qPCR analysis revealed elevated <i>Prss56</i> mRNA levels in <i>Prss56</i>^<i>F/F</i><i>; Ubc-Cre</i>^<i>ERT2</i> retina compared to <i>Prss56</i>^<i>F/+</i><i>; Ubc-Cre</i>^<i>ERT2</i> retina following tamoxifen injection at P8, P13, or P18 (shown are data from mice harvested at P17 and P60, respectively). (<b>M</b>) Following <i>Prss56</i> ablation at P18, the eyes display a hyperopic shift in refraction compared to control eyes at 3 months (N = 6 per group). Values are presented as mean ± SD (or mean ± SEM in <b>L</b>); * p<0.05, ** p<0.01, *** p<0.001, t-test.