# Zinc regulation of lipidome remodeling during boar sperm capacitation

## TL;DR

Zinc ions influence lipid changes in boar sperm during capacitation, a process needed for fertilization, by stabilizing certain membrane lipids and altering others.

## Contribution

The study reveals zinc's specific role in lipidome remodeling during sperm capacitation, identifying distinct classes of lipid responses.

## Key findings

- Zinc inhibits or stabilizes 12 out of 30 capacitation-associated lipid shifts.
- Zinc enhances capacitation-linked lipid changes in 2 lipid species.
- Lipids are categorized into four zinc-response types based on their behavior during capacitation.

## Abstract

Sperm capacitation is essential for fertilization and is characterized by a cascade of biochemical signaling and membrane remodeling events. This process is highly dependent on membrane composition. Profiling lipid alterations provides a critical window into the molecular underpinnings of capacitation and the regulatory influence of zinc ions (Zn2+). Metabolomic studies in boar sperm have shown that capacitation coincides with broad shifts in metabolite abundance and that extracellular zinc attenuates or redirects many of these changes, highlighting its role as a key modulator. To extend this framework to the lipidome, we profiled boar sperm under three conditions: non-capacitated (0 h), capacitated in vitro (4 h), and capacitated with extracellular zinc (4 h + Zn), using liquid chromatography–mass spectrometry and image-based flow cytometry to validate capacitation status. Relative to 0 h, capacitation was associated with altered abundances of 30 lipids (P < 0.05) spanning several lipid categories: fatty acyls (n = 8), sterol lipids (n = 7), sphingolipids (n = 1), glycerolipids (n = 3), glycerophospholipids (n = 4), and unannotated lipids (n = 9). When exogenous Zn2+ was supplemented during in vitro capacitation, 12 of these shifts were maintained at 0 h-like levels (P < 0.05), suggesting an inhibiting or stabilizing role. In 2 of 16 hits, exogenously supplemented Zn2+ enhanced the capacitation-associated change (P < 0.05), whereas in the remaining 14 it exerted no measurable effect (P > 0.05). When exogenous Zn2+ was supplemented during in vitro capacitation, distinct lipid shifts were identified and organized using Tukey’s lipid-pattern classification based on significance (P < 0.05) and directionality, organizing them into four categories: Type-1 lipids (capacitation-associated), Type-2 lipids (zinc-inhibited), Type-3 lipids (zinc-specific response), and Type-4 lipids (zinc-enhanced). These categories describe distinct modes of lipid regulation, where some species remained unaffected by zinc (Type-1, n = 16), others were stabilized or inhibited from progressing toward capacitation-associated levels (Type-2, n = 12), a subset responded exclusively to zinc independent of capacitation (Type-3, n = 4), and a small group exhibited amplified capacitation-linked shifts under zinc supplementation (Type-4, n = 2). Together, these data reveal a class-specific, zinc-dependent architecture of lipid remodeling that integrates metabolic and membrane regulation within the broader capacitation cascade.

Graphical AbstractGraphical Abstract Lipid species responses to in vitro capacitation with and without exogenous zinc. (a) Schematic representation of the sperm lipidome under non-capacitated conditions (0 h). The red sperm depicts uncapacitated spermatozoa, with the associated lipidome labeled as the non-capacitated sperm lipidome. Colored lipid symbols represent lipid species that were subsequently classified in panel (b) and are shown by lipid class as follows: fatty acyls (FA; light blue), glycerophospholipids (GP; dark blue), sterol lipids (SL; green), glycerolipids (GL; orange), and unannotated lipids (UA; purple). Lipid class abbreviations are embedded within each symbol to ensure clear identification independent of color. The distribution of colored symbols is representative and does not indicate absolute abundance. (b) Lipid response pattern classifications summarizing the distribution of differentially abundant lipids by zinc-response type, defined using Tukey’s honestly significant difference post hoc comparisons across 0 h, 4 h, and 4 h + Zn conditions. Type-1 lipids are zinc-uninhibited (16 of 34, purple). Type-2 lipids are zinc-inhibited (12 of 34, blue), with complete inhibition shown in light blue and partial inhibition in dark blue. Type-3 lipids are zinc-modulated and capacitation-independent (4 of 34, green). Type-4 lipids are zinc-enhanced (2 of 34, orange). Created in BioRender. Kerns, K. (2025) https://BioRender.com/2i6406v

Graphical Abstract Lipid species responses to in vitro capacitation with and without exogenous zinc. (a) Schematic representation of the sperm lipidome under non-capacitated conditions (0 h). The red sperm depicts uncapacitated spermatozoa, with the associated lipidome labeled as the non-capacitated sperm lipidome. Colored lipid symbols represent lipid species that were subsequently classified in panel (b) and are shown by lipid class as follows: fatty acyls (FA; light blue), glycerophospholipids (GP; dark blue), sterol lipids (SL; green), glycerolipids (GL; orange), and unannotated lipids (UA; purple). Lipid class abbreviations are embedded within each symbol to ensure clear identification independent of color. The distribution of colored symbols is representative and does not indicate absolute abundance. (b) Lipid response pattern classifications summarizing the distribution of differentially abundant lipids by zinc-response type, defined using Tukey’s honestly significant difference post hoc comparisons across 0 h, 4 h, and 4 h + Zn conditions. Type-1 lipids are zinc-uninhibited (16 of 34, purple). Type-2 lipids are zinc-inhibited (12 of 34, blue), with complete inhibition shown in light blue and partial inhibition in dark blue. Type-3 lipids are zinc-modulated and capacitation-independent (4 of 34, green). Type-4 lipids are zinc-enhanced (2 of 34, orange). Created in BioRender. Kerns, K. (2025) https://BioRender.com/2i6406v

Zinc fine-tunes lipid remodeling during boar sperm capacitation, stabilizing sterol-rich membrane regions while selectively reprogramming fatty-acid dynamics. These findings reveal zinc as a key regulator linking sperm membrane composition to fertilization readiness.