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Overview and major findings

Zhang et al. investigate how mechanical forces are transduced during labor (parturition) via the mechanosensitive ion channels PIEZO1 and PIEZO2. Using mouse genetic models, the authors demonstrate that deletion of Piezo1 and Piezo2 in tissues caudal to the diaphragm (Hoxb8^Cre, targets both sensory neuronal and non-neuronal cells) results in weakened uterine contractions and parturition defects. To dissect the contribution of specific tissues, the authors then utilized tissue-specific genetic deletion models, which showed that Piezo1/2 double knockout in uterine smooth muscle tissues or uterine-innervating sensory neurons each produces modest labor defects, but combined deletion causes severe labor impairment.  At the molecular level, the authors utilized single-nucleus sequencing in double knockout Hoxb8^Cre mice to reveal a downstream relationship that mechanistically links deficient mechanotransduction to impaired uterine contractile biology through downregulation of smooth muscle Gja1, which encodes the gap junction protein connexin 43—a known key player in uterine contractility that is expressed in the mouse and human myometrium and was previously shown to be critical for normal parturition. The authors propose a model where uterine smooth muscle senses mechanical stretch/pressure via PIEZO1, while sensory nerves detect stretch via PIEZO2, which together coordinate effective labor. They suggest that mechanotransduction through PIEZO channels is a critical but under‐appreciated element of parturition and note that other (PIEZO-independent) mechanotransduction mechanisms may be at play. While prior work has also looked at mechanosensitive channels in the myometrium (PMID: 35941750), linking mechanosensory ion channels directly to parturition and uterine contraction coordination is novel. The current study adds an important piece, which demonstrates that uterine mechanical force is sensed and translated into molecular changes that support coordinated contraction downstream. These data also have potentially promising impact for human labor dysfunction, especially since, as demonstrated by the authors, expression pattern of PIEZO channels in human tissues during term pregnancy closely aligns with that in mouse tissues.

Major comments

• No major comments.

Minor comments

• It would be helpful to include the appropriate negative controls for the smFISH experiments, e.g., Fig. 1A & C and Fig. S1B, E, F, & H, in the main figures or supplemental materials to clarify background signal and ensure specificity of the probes.

• In Fig. 1E–F/page 3 (line 19–24), light-sheet imaging shows that Piezo2⁺ sensory fibers primarily innervate the lower reproductive tract. To provide a more complete picture, it would be helpful to state where else was labeling observed using this AAV-mediated approach, and whether any of these labeled tissues may directly/indirectly contribute to uterine contractility.

• For Fig. S1L, it would be helpful to include separate single-channel panels alongside the merged image to more clearly show the lack of overlap between CTB⁺ and Sst⁺ cells.

• On page 3 (line 36–37), a brief statement clarifying how the referenced Sun1 reporter system works (e.g., add “a Cre-dependent fluorescent reporter which labels the inner nuclear membrane”) would aid readers who may be less familiar with this tool.

• In Fig. 2 legend/page 16 (line 2–3), the statement that “HoxB8-Cre is used to knock out Piezo1 and Piezo2 in most cells relevant to pregnancy” may overstate the breadth of the model. Since some pregnancy-relevant tissues lie outside the HoxB8 domain (e.g., pituitary and hypothalamus, which control neuroendocrine signals that initiate and amplify uterine contractions), it may be helpful to rephrase this for accuracy. For example, the authors may consider clearly stating that “all uterine cell types apart from the epithelium are targeted by this Cre line”.

• On page 3 (line 40–41), there appears to be a typographical error: “HoxB8^Cre; Piezo2^f/f; Piezo2^f/f, referred to as HoxB8; P1; P2” should read “HoxB8^Cre; Piezo1^f/f; Piezo2^f/f, referred to as HoxB8; P1; P2” to aid readability.

• On page 4 (line 35–37), the text notes that all controls deliver their first pup before ZT3 whereas roughly half of the knockout animals deliver after this time point, and indicates that ZT3 was hence chosen as the reference point throughout the paper. It would be helpful to readers if the authors briefly comment on the potential broader biological significance of this specific time point. For example, is anything known about circadian control over labor initiation time, and could this be related to mechanosensation?

• On page 4 (line 42), it would be helpful to clarify in the main text or methods section what numerical or analytical threshold was used to distinguish “high-pressure peaks” from lower pressure events so that readers can contextualize the biological significance of this analysis.

• On page 6 (line 28–30), it would strengthen the statement “…As expected for PIEZOs as ion channels that influence transcription indirectly through cumulative cation influx…” to include a citation to prior work supporting this mechanism.

• In Fig. S3A, it appears that the first and fifth control dot plot patterns are identical to the first and fifth KO plots (respectively). Similarly, the second, sixth, and ninth control plots appear nearly identical to each other as well as to those in the KO group, essentially showing a single contraction of the same strength. Could the authors clarify whether this is an error in data plotting/labeling or simply coincidence?

• While HoxB8-Cre only targets tissues caudal to the diaphragm, PGR-Cre is also expressed in the pituitary gland (see relevant JAX labs mouse line webpage). Did the authors perform any controls to ensure that loss of PIEZO channels in these tissues does not contribute to the observed labor phenotypes? This is particularly relevant because pituitary-derived oxytocin is a major regulator of uterine contractions, contraction intensity, and cervical dilation, and is even used clinically to induce or augment labor.

Competing interests

The authors declare that they have no competing interests.

Use of Artificial Intelligence (AI)

The authors declare that they did not use generative AI to come up with new ideas for their review.