Proliferating cells known as neoblasts include pluripotent stem cells (PSCs) that sustain tissue homeostasis and regeneration of lost body parts in planarians. However, the lack of markers to prospectively identify and isolate these adult PSCs has significantly hampered their characterization. We used single-cell RNA sequencing (scRNA-seq) and single-cell transplantation to address this long-standing issue. Large-scale scRNA-seq of sorted neoblasts unveiled a novel subtype of neoblast (Nb2) characterized by high levels of PIWI-1 mRNA and protein and marked by a conserved cell-surface protein-coding gene, tetraspanin 1 (tspan-1). tspan-1-positive cells survived sub-lethal irradiation, underwent clonal expansion to repopulate whole animals, and when purified with an anti-TSPAN-1 antibody, rescued the viability of lethally irradiated animals after single-cell transplantation. The first prospective isolation of an adult PSC bridges a conceptual dichotomy between functionally and molecularly defined neoblasts, shedding light on mechanisms governing in vivo pluripotency and a source of regeneration in animals. VIDEO ABSTRACT.

Aggregation of damaged or misfolded proteins is a protective mechanism against proteotoxic stress, abnormalities of which underlie many aging-related diseases. Here, we show that in asymmetrically dividing yeast cells, aggregation of cytosolic misfolded proteins does not occur spontaneously but requires new polypeptide synthesis and is restricted to the surface of ER, which harbors the majority of active translation sites. Protein aggregates formed on ER are frequently also associated with or are later captured by mitochondria, greatly constraining aggregate mobility. During mitosis, aggregates are tethered to well-anchored maternal mitochondria, whereas mitochondria acquired by the bud are largely free of aggregates. Disruption of aggregate-mitochondria association resulted in increased mobility and leakage of mother-accumulated aggregates into the bud. Cells with advanced replicative age exhibit gradual decline of aggregates-mitochondria association, likely contributing to their diminished ability to rejuvenate through asymmetric cell division.

Among vertebrates, only microchiropteran bats, cetaceans and some rodents are known to produce and detect ultrasounds (frequencies greater than 20 kHz) for the purpose of communication and/or echolocation, suggesting that this capacity might be restricted to mammals. Amphibians, reptiles and most birds generally have limited hearing capacity, with the ability to detect and produce sounds below approximately 12 kHz. Here we report evidence of ultrasonic communication in an amphibian, the concave-eared torrent frog (Amolops tormotus) from Huangshan Hot Springs, China. Males of A. tormotus produce diverse bird-like melodic calls with pronounced frequency modulations that often contain spectral energy in the ultrasonic range. To determine whether A. tormotus communicates using ultrasound to avoid masking by the wideband background noise of local fast-flowing streams, or whether the ultrasound is simply a by-product of the sound-production mechanism, we conducted acoustic playback experiments in the frogs' natural habitat. We found that the audible as well as the ultrasonic components of an A. tormotus call can evoke male vocal responses. Electrophysiological recordings from the auditory midbrain confirmed the ultrasonic hearing capacity of these frogs and that of a sympatric species facing similar environmental constraints. This extraordinary upward extension into the ultrasonic range of both the harmonic content of the advertisement calls and the frog's hearing sensitivity is likely to have co-evolved in response to the intense, predominantly low-frequency ambient noise from local streams. Because amphibians are a distinct evolutionary lineage from microchiropterans and cetaceans (which have evolved ultrasonic hearing to minimize congestion in the frequency bands used for sound communication and to increase hunting efficacy in darkness), ultrasonic perception in these animals represents a new example of independent evolution.

Transplantation of hematopoietic stem cells (HSCs) from human umbilical cord blood (hUCB) holds great promise for treating a broad spectrum of hematological disorders including cancer. However, the limited number of HSCs in a single hUCB unit restricts its widespread use. Although extensive efforts have led to multiple methods for ex vivo expansion of human HSCs by targeting single molecules or pathways, it remains unknown whether it is possible to simultaneously manipulate the large number of targets essential for stem cell self-renewal. Recent studies indicate that N-methyladenosine (mA) modulates the expression of a group of mRNAs critical for stem cell-fate determination by influencing their stability. Among several mA readers, YTHDF2 is recognized as promoting targeted mRNA decay. However, the physiological functions of YTHDF2 in adult stem cells are unknown. Here we show that following the conditional knockout (KO) of mouse Ythdf2 the numbers of functional HSC were increased without skewing lineage differentiation or leading to hematopoietic malignancies. Furthermore, knockdown (KD) of human YTHDF2 led to more than a 10-fold increase in the ex vivo expansion of hUCB HSCs, a fivefold increase in colony-forming units (CFUs), and more than an eightfold increase in functional hUCB HSCs in the secondary serial of a limiting dilution transplantation assay. Mapping of mA in RNAs from mouse hematopoietic stem and progenitor cells (HSPCs) as well as from hUCB HSCs revealed its enrichment in mRNAs encoding transcription factors critical for stem cell self-renewal. These mA-marked mRNAs were recognized by Ythdf2 and underwent decay. In Ythdf2 KO HSPCs and YTHDF2 KD hUCB HSCs, these mRNAs were stabilized, facilitating HSC expansion. Knocking down one of YTHDF2's key targets, Tal1 mRNA, partially rescued the phenotype. Our study provides the first demonstration of the function of YTHDF2 in adult stem cell maintenance and identifies its important role in regulating HSC ex vivo expansion by regulating the stability of multiple mRNAs critical for HSC self-renewal, thus identifying potential for future clinical applications.