Sonic hedgehog (Shh), which regulates proliferation in many contexts, functions as a limb morphogen to specify a distinct pattern of digits. How Shh's effects on cell number relate to its role in specifying digit identity is unclear. Deleting the mouse Shh gene at different times using a conditional Cre line, we find that Shh functions to control limb development in two phases: a very transient, early patterning phase regulating digit identity, and an extended growth-promoting phase during which the digit precursor mesenchyme expands and becomes recruited into condensing digit primordia. Our analysis reveals an unexpected alternating anterior-posterior sequence of normal mammalian digit formation. The progressive loss of digits upon successively earlier Shh removal mirrors this alternating sequence and highlights Shh's role in cell expansion to produce the normal digit complement.

The development of photocaging groups activated by near-IR light would enable new approaches for basic research and allow for spatial and temporal control of drug delivery. Here we report a near-IR light-initiated uncaging reaction sequence based on readily synthesized C4′-dialkylamine-substituted heptamethine cyanines. Phenol-containing small molecules are uncaged through sequential release of the C4′-amine and intramolecular cyclization. The release sequence is initiated by a previously unexploited photochemical reaction of the cyanine fluorophore scaffold. The uncaging process is compatible with biological milieu and is initiated with low intensity 690 nm light. We show that cell viability can be inhibited through light-dependent release of the estrogen receptor antagonist, 4-hydroxycyclofen. In addition, through uncaging of the same compound, gene expression is controlled with near-IR light in a ligand-dependent CreERT/LoxP-reporter cell line derived from transgenic mice. These studies provide a chemical foundation that we expect will enable specific delivery of small molecules using cytocompatible, tissue penetrant near-IR light.

e TAF7, a component of the TFIID complex that nucleates the assembly of transcription preinitiation complexes, also independently interacts with and regulates the enzymatic activities of other transcription factors, including P-TEFb, TFIIH, and CIITA, ensuring an orderly progression in transcription initiation. Since not all TAFs are required in terminally differentiated cells, we examined the essentiality of TAF7 in cells at different developmental stages in vivo. Germ line disruption of the TAF7 gene is embryonic lethal between 3.5 and 5.5 days postcoitus. Mouse embryonic fibroblasts with TAF7 deleted cease transcription globally and stop proliferating. In contrast, whereas TAF7 is essential for the differentiation and proliferation of immature thymocytes, it is not required for subsequent, proliferation-independent differentiation of lineage committed thymocytes or for their egress into the periphery. TAF7 deletion in peripheral CD4 T cells affects only a small number of transcripts. However, T cells with TAF7 deleted are not able to undergo activation and expansion in response to antigenic stimuli. These findings suggest that TAF7 is essential for proliferation but not for proliferation-independent differentiation.