Male fertility fundamentally relies on the precisely orchestrated process of spermatogenesis, which governs the continuous generation of spermatozoa [1]. This developmental cascade initiates with spermatogonial differentiation into primary spermatocytes, progressing through meiotic recombination to produce haploid spermatids [2], [3]. Subsequent spermiogenesis involves dynamic cellular remodeling, including nuclear elongation, chromatin condensation, acrosomal vesicle formation, cytoskeletal reorganization, and cytoplasmic extrusion—key processes essential for generating fertilization-competent sperm [4]. The genomic complexity underlying testicular function is evidenced by transcriptional activation of nearly 16,500 protein-coding genes, representing over 80% of mammalian coding capacity [5], [6]. While targeted mutagenesis studies have functionally characterized numerous testis-enriched genes, with many demonstrating spermatogenic arrest phenotypes, substantial knowledge gaps persist regarding the biological significance of male germ cell-specific genetic elements [7].

Leucine-rich repeat and WD40 repeat domain-containing protein 1 (LRWD1), also known as origin recognition complex-associated protein (ORCA), forms a complex with the origin recognition complex (ORC) to stabilize its binding to replication origins on chromatin, featuring an N-terminal leucine-rich repeat (LRR) domain and three C-terminal WD40 repeats critical for chromatin interactions [8], [9]. Previous studies have shown that LRWD1 interacts with repressive histone modifications marking heterochromatin, contributing to the silencing of major satellite repeats in mouse pericentric heterochromatin and mediating the establishment and maintenance of H3K9me3 enrichment in specific genomic regions in human cells[10], [11], [12]. Additionally, LRWD1 is crucial for heterochromatin organization and the assembly of the pre-replication complex (pre-RC) during the G1 phase in somatic cells[13], [14]. Germline expression profiling identifies LRWD1 in testicular tissues with dynamic subcellular localization-cytoplasmic during spermatogenic differentiation and neck-associated in mature spermatozoa [15], [16], [17]. Despite these observations suggesting potential roles in heterochromatin organization, replication dynamics, and survival of developing sperm cells, the functional necessity of LRWD1 in mammalian spermatogenesis remains undefined.

Here, we showed that Lrwd1 exhibits evolutionary conservation across eutherians and demonstrates predominant testicular expression. Through generation of germ cell-specific Lrwd1 knockout mice using Stra8-Cre (Lrwd1-sKO), we systematically evaluated reproductive parameters. Notably, Lrwd1-sKO males maintained normal fertility indices, showing preserved spermatogenic progression, unaltered sperm output, and intact sperm motility characteristics. These findings suggest that, despite its high expression in the testis, Lrwd1 is dispensable for fertility in mice.