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Molecular Mechanisms of Milk Production Reveal an Evolutionary Back-up Plan

    Rank proteins, mammary glands, luminal cells, basal receptors, basal cells, milk production

    Written by: Lauren Milligan Newmark, Ph.D. | Issue # 118 | 2024

    • To understand the influence of the Rankl protein on lactation, researchers turned off cellular receptors for Rankl in two types of cells within a mouse mammary gland.
    • Receptor loss in basal cells did not affect lactation performance, but receptor loss in luminal cells led to lactation failure for the first litter.
    • Mice with receptor loss on luminal cells were able to successfully lactate with their second and third litters due to novel finding that basal stem cells could produce Rank-positive luminal cells.

    If you have ever wondered what one teaspoon of baking soda does for an entire batch of cookies, omit this ingredient the next time you bake, and your flat cookies will clue you in. Biologists actually use the “omit one ingredient” approach when trying to understand the function of a gene or protein in an organism. Sometimes the best way to figure out what something does is to take it away and see what happens. 

    In a new study [1], a team of Spanish researchers applied this method to understand how one protein affects milk production by the mammary gland [1]. The protein of interest is called Rankl (Receptor Activator of Nuclear factor-Kappa beta Ligand). The research team knew that the expression of Rankl within the mammary gland was necessary for proper mammary gland development and milk production based on previous animal studies that turned off Rankl signaling in all cells. Hoping to zoom in on the specific impact of this protein on lactation, the Spanish team turned off the Rankl signaling pathway in just two cellular lineages.

    The team focused on luminal cells and basal cells, the two types of mammary epithelial cells [1]. During embryonic development, basal stem cells give rise to luminal and basal cell lines. During puberty, these cell types are still in their progenitor phase and have yet to differentiate or specialize. However, by the onset of lactation, luminal cells give rise to alveolar and ductal cells (milk secreting cells), whereas basal cells differentiate into myoepithelial cells (which are involved in milk ejection) [2, 3]. 

    Using a mouse model, the researchers blocked the signal from the Rankl protein by genetically removing (aka “knocking out”) the instructions for making the Rankl cell surface receptor on either luminal or basal cells. Then, they compared tissue development and lactation performance between mice with basal Rank receptor loss, mice with luminal Rank receptor loss, and control mice (no genetic interventions). 

    Mice with basal Rank receptor loss did not exhibit developmental changes or issues with lactation performance over multiple pregnancies. The loss of basal Rankl signaling did not appear to impact the ability of basal cells to differentiate over the course of the female mouse’s reproductive lifespan or the function of the mammary gland during pregnancy or lactation [1]. Rankl protein, it seems, was not integral to basal cell function.

    In contrast, the team found that mice with luminal Rank receptor loss had fewer luminal progenitor cells during puberty compared with control mice. During pregnancy, they observed an impaired response of luminal cells to the hormones progesterone and prolactin, which led to abnormal differentiation of alveolar cells [1]. These aberrant alveolar cells were unable to secrete milk, resulting in lactation failure.

    If the research study ended there, it would be interesting and scientifically valuable—the research team was able to establish that Rankl proteins are required for successful lactation because of their influence on luminal cell progenitor populations and luminal cell differentiation into alveolar cells [1]. But the researchers decided to follow their knockout mice through multiple pregnancies and, in doing so, were able to make an even more ground-breaking finding. 

    To the researchers’ surprise, luminal rank loss mice who couldn’t produce milk for their first litter were able to do so for all the pups in their second litter and most pups in their third litter [1]. Unlike the first pregnancy, alveolar cell development was normal during the second and third pregnancy, resulting in what the researchers describe as “an almost normal lactating gland” [1]. What had happened between the first and second pregnancies to bring about this functional change? Amazingly, Rank-deleted luminal cells had been replaced during the second pregnancy by Rank-positive cells [1]. And even more remarkable, the cellular grandparent of these functional luminal cells were basal cells. 

    Basal cells in the mammary epithelium were known to be bipotent—able to produce two cell types—during embryonic development but not in adulthood. The researchers hypothesize that in their mouse model, basal cell bipotency was triggered by molecular changes brought about by lactation failure specifically in the context of luminal Rank loss [1]. Exactly which molecular changes trigger basal cells to come to the functional rescue of the abnormal luminal cells is still unknown, but the study authors hypothesize it was the combination of inflammatory signals generated by the Rank-deleted luminal progenitors and/or the abnormal alveolar cells they produced and an excess of Rankl protein, the result of fewer receptors for Rankl protein binding [1]. 

    It is tempting to propose that the triggering of basal cell bipotency is an evolved mechanism to protect lactation in mice, and potentially in other mammals (even humans). If normal alveolar cell development is indeed critical to successful lactation, having a back-up plan to prevent lactation failure in second pregnancies and beyond would ensure reproductive success across mammals. 


    1. Rocha AS, Collado-Solé A, Graña-Castro O, Redondo-Pedraza J, Soria-Alcaide G, Cordero A, Santamaría PG, González-Suárez E. Luminal Rank loss decreases cell fitness leading to basal cell bipotency in parous mammary glands. Nature Communications. 2023 Oct 9;14(1): 6213.
    2. Tiede B, Kang Y. From milk to malignancy: the role of mammary stem cells in development, pregnancy and breast cancer. Cell Research. 2011 Feb;21(2): 245-57.
    3. Cristea S, Polyak K. Dissecting the mammary gland one cell at a time. Nature Communications. 2018 Jun 26;9(1): 2473.